WO2023218226A1 - Procédé de croissance d'un biomatériau sur un substrat à l'aide d'un système de chevron flottant - Google Patents

Procédé de croissance d'un biomatériau sur un substrat à l'aide d'un système de chevron flottant Download PDF

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Publication number
WO2023218226A1
WO2023218226A1 PCT/IB2022/054415 IB2022054415W WO2023218226A1 WO 2023218226 A1 WO2023218226 A1 WO 2023218226A1 IB 2022054415 W IB2022054415 W IB 2022054415W WO 2023218226 A1 WO2023218226 A1 WO 2023218226A1
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Prior art keywords
floating
substrate
further characterized
biomaterial
rafting
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PCT/IB2022/054415
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English (en)
Inventor
Axel Alejandro GÓMEZ-ORTIGOZA AGUIRRE
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Polybion S.L.
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Priority to PCT/IB2022/054415 priority Critical patent/WO2023218226A1/fr
Publication of WO2023218226A1 publication Critical patent/WO2023218226A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P19/00—Preparation of compounds containing saccharide radicals
    • C12P19/04—Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/02—Acetobacter

Definitions

  • the present invention relates to a method for growing a biomaterial directly formed and/or secreted onto a substrate using a floating rafting system over a liquid culturing medium for incubating said biomaterial, the biomaterial obtained with said method, and a floating rafting system used for performing said method.
  • plant-based materials which are obtained from different organic waste, for example, from apple, pineapple, mango, cactus, grapes, among others.
  • Other sources of organic material can be animal cells from mammalian, reptile, fish, birds, and amphibians, among others. Fungi and bacteria can also be used for treating organic raw materials for the manufacturing of everyday items.
  • Document WO 2003/105726 refers to a composite prosthetic implant which comprises a textile support of which at least a portion of the surface is covered by a lyophilisate of a biocompatible material.
  • This document discloses a method for manufacturing a composite prosthetic implant onto a textile support impregnated with a biocompatible material, wherein the biocompatible material can be selected from one of hyaluronic acid, alginates, polypeptides and polycaprolactone, mixtures and derivatives thereof.
  • this patent document discloses that there is a pouring process of a second biocompatible material onto a textile support.
  • This document refers to a process for impregnating a textile support with a biocompatible material to manufacture a final product.
  • this document does not refer to the use of a floating rafting system submerged into a culture liquid medium to obtain the biomaterial.
  • WO 2019/198002 discloses a method of manufacturing artificial leather, wherein a textile support is prepared, preferably with a base made of synthetic microfiber nonwoven fabric, with a polyurethane coagulate and applying color coats, wherein said prepared textile support is embossed to obtain a product.
  • a non-woven fabric as substrate, but this document does not consider the use of a floating rafting system submerged into a liquid culturing medium used to place the textile (substrate) on which the biomaterial will grow.
  • W02020037486A1 discloses a process that includes (i) first, contacting a textile with an aqueous solution containing a cationic hydroxyethylcellulose polymer to form a modified textile component; (ii) subsequently, impregnating the modified textile component with an aqueous polyurethane dispersion externally stabilized with an anionic surfactant; and (iii) precipitating the polyurethane in the modified textile component.
  • This document anticipates the impregnation of a modified textile with an aqueous polyurethane dispersion to form a synthetic leather, where the textile is modified by an aqueous solution containing a cationic hydroxyethylcellulose polymer.
  • the document does not disclose the use of a floating rafting system for placing the textile onto which the biomaterial will grow.
  • US Patent 9,840,745 B2 discloses a method of manufacturing a product, wherein a container apparatus including a culture medium and a support element therein is provided in which at least one reinforcing structure on the support element is placed. Dermal cells are introduced into the culture medium so that the dermal cells grow and divide to form a tissue layer including a first stratum on a first side of the tissue layer, a second stratum on a second side of the tissue layer, and a third stratum disposed between the first stratum and the second stratum.
  • the process described in this document considers the use of a container apparatus which includes a culture medium and a support element therein. However, the document does not disclose the use of a floating rafting system for placing the textile onto which the biomaterial will grow.
  • EP 3 337 923 B1 describes engineered, reinforced leather materials (engineered leathers) including a composite of a fibrous scaffold and a collagen network formed by cultured cells (e.g., fibroblasts). These composites are tanned to stabilize the collagen network and interactions between the fibrous scaffold and collagen network. These engineered leathers may be referred to as fiber-reinforced biological tissue composites.
  • the document also describes a method of forming a fiber-reinforced biological tissue composite by culturing tissue-producing cells (e.g., fibroblasts) on a fibrous scaffold having a functional group selected from the group consisting of amine (-NH2), carboxylic acid (-COOH), sulfhydryl (-SH), and hydroxyl (-OH), and combinations thereof, which may be cross-linked (e.g., by tanning) to the tissue and/or proteins, such as collagen, formed and/or secreted by the cells.
  • Tanning may be performed after the scaffold fibers have been at least partially covered in cultured cells and the extracellular matrix released by the cultured cells.
  • the document does not disclose the use of a floating rafting system for placing the textile onto which the biomaterial will grow.
  • Some of the mentioned methods are useful for obtaining a biomaterial which in a subsequent step will be added to a support so that an additional laminating step is required by using a glue or adhesive onto the substrate.
  • microorganism is dependent on the presence of water.
  • the substances from which microorganisms synthesize their cell material and obtain their energy and nutrients are dissolved in water.
  • Different microorganisms have very different requirements for the composition of nutrient media as well as for other environmental conditions. Because of this, numerous recipes for the composition of microbiological media have been devised. Basically, all nutrient media must fulfill with some minimal requirements e.g., they must supply as utilizable compounds all elements that take part in the synthesis of the cell substance (“General Microbiology”, G. Schlegel, C. Zaborosch, 7 th ed., page 193).
  • Bacterial cellulose is an organic compound with the formula (CeHioO)n produced by certain types of bacteria. While cellulose is a basic structural material of most plants, it is also produced by bacteria, principally of the genera Acetobacter, Sarcina ventriculi and Agrobacterium. Bacterial, or microbial, cellulose has different properties from plant cellulose and is characterized by high purity, strength, moldability and increased water holding ability. (Jonas, R.; Farah, Luiz F. (1998). "Production and application of microbial cellulose” Polymer Degradation and Stability, vol.59 (Issues 1-3): pages 101-106).
  • the present invention solves the disadvantages of the prior art by providing a method for growing biomaterial directly onto substrates by using a floating rafting system avoiding the use of the additional step of laminating the sheet of biomaterial for adhering the substrate to the biomaterial.
  • the method of the present invention also eliminates the use of glues or adhesives, reducing the footprint of the material, increasing the bonding strength, and overall increasing the process efficiency (energetic, ecological footprint, cost, cycle time, etc.).
  • the present invention comprises the use of a liquid culturing medium enriched with cells.
  • Another object of the invention is to eliminate the use of glues, adhesives, and the like, reducing the footprint of the material, and increasing the bonding strength between the biomaterial and the substrate.
  • Another object of the invention is to provide an organic material which helps to diminish the environmental impact improving the energetic efficiency, ecological footprint, costs, cycle time, etc.
  • One additional object of the present invention is to provide a floating rafting system which comprises a specific structure for growing a biomaterial onto the substrate.
  • the floating rafting system comprises a frame and a substrate.
  • the floating rafting system comprises fastening means for fastening a substrate onto the frame.
  • the floating rafting system comprises a supporting mesh on which the substrate can be superposed.
  • the frame being a rigid structure that comprises profiles made of a material selected from any of standard aluminum and its alloys, e.g., AICu, AlSi, AIMg, AlZn, AIMn, AITi, AINi, among others; plastic, e.g., polyvinyl chloride (PVC), polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET), polyethylene (PE), polypropylene (PP), expanded polystyrene (EPS), among others; wood or cork, or any other buoyant material which prevents the frame to sink due to the light weight of the material when the structure is placed onto a liquid medium, wherein optionally floaters are attached to said profiles, to increase and/or control the buoyancy of the system.
  • plastic e.g., polyvinyl chloride (PVC), polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene ter
  • the floating rafting system comprises a supporting mesh attached to the frame.
  • the supporting mesh can be made of any suitable rigid material such as plastic or metal.
  • the supporting mesh can have a sieve mesh size suitable not only to allow the liquid culturing medium to pass through it, but to superpose the substrate evenly tensed thereon, when necessary.
  • the sieve mesh size can be of 10 mm or less.
  • a substrate which can be a textile selected from a woven or non-woven textile, an electronic circuit, silicon circuit board, conductive thread, wire or similar elements, is also provided to be fastened directly to the frame of the floating rafting system using the fastening means, wherein in an embodiment the fastening means in the floating rafting system are fastening lanes.
  • the substrate is fastened to the frame in such a manner that it is completely and evenly tensed, in order to ensure an even biomaterial growth along the substrate.
  • This can be achieved, for example, by using a standard vinyl spline (cord) inserted into the fastening lane, fastening the substrate therebetween.
  • the substrate can be superposed on the supporting mesh without the need to be fasted directly to the frame.
  • a woven or non-woven textile or other suitable substrate can be placed evenly tensed over the supporting mesh without the need to be fastened directly to the floating rafting system because the tension of the supporting mesh is transferred to the textile substrate.
  • the substrate can consist of loose fibers widespread over the supporting mesh.
  • the loose fibers can be selected from plastic fibers e.g., polyvinyl chloride (PVC), polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET), polyethylene (PE), polypropylene (PP), expanded polystyrene (EPS), Kevlar fibers, carbon fibers, cellulose fibers, lignocellulose fibers, among others, and combinations thereof.
  • plastic fibers e.g., polyvinyl chloride (PVC), polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET), polyethylene (PE), polypropylene (PP), expanded polystyrene (EPS), Kevlar fibers, carbon fibers, cellulose fibers, lignocellulose fibers, among others, and combinations thereof.
  • plastic fibers e.g., polyvinyl chloride (PVC), polylactic acid (PL
  • the culturing medium can comprise any type of cells desired to be grown, such as mammalian cells, fish cells, amphibian cells, insect cells, plant cells, bacterial cells and fungal cells, including yeast cells and mold cells.
  • the liquid culturing medium can be a bacterial cellulose culturing medium, comprises dried food waste, dried black tea and water.
  • the bacterium in the bacterial culturing medium mainly comprises acetobacter xylinum.
  • the floating rafting system is left floating into the liquid culturing medium for several days until a homogeneous layer of biomaterial has grown evenly onto the substrate.
  • the floating rafting system is flipped 180° and is left floating into the liquid culturing medium for another growth period, so there is an even growth of biomaterial throughout both sides of the substrate.
  • the floating rafting system is removed from the liquid culturing medium.
  • the floating rafting system is removed from the liquid culturing medium, the substrate with the biomaterial is separated from the floating rafting system.
  • the fastening means are separated from the frame thus releasing the wet substrate with the biomaterial that has grown over the time. Since the biomaterial has grown directly onto the substrate, there is no chemical crosslinking between the biomaterial and the substrate, thus the need for a lamination process is prevented.
  • the floating rafting system is set to dry before the substrate is removed.
  • the biomaterial After the biomaterial is extracted, it could be subjected to a post-processing step in order to produce a commercial product.
  • the most preferable post-processing process is tanning.
  • the floating rafting system especially designed to be used in the method for growing a biomaterial onto a substrate can be used over, representing an economic and ecological system.
  • Figure 1 is a perspective view showing a floating rafting system in a liquid culturing medium according to an embodiment of the present invention.
  • Figure 2 shows a perspective view of a preferred embodiment of the floating rafting system.
  • Figure 3 is an exploded view showing the floating rafting system with the substrate.
  • Figure 4 is a bottom view of a preferred floating rafting system.
  • Figure 5 is a perspective view of the floating rafting system showing the biomaterial grown onto the floating rafting system.
  • the invention generally relates the method to grow a biomaterial formed and/or secreted by cells onto a substrate by using a rafting floating system placed over a liquid culturing medium; wherein said biomaterial includes at least one polysaccharide, lipid or protein, either separately or a combination of the same.
  • a rafting floating system (1 comprises a frame (2), fastening means (4) (see Figure 4), and a substrate (5) fastened to the frame (2) through the fastening means (4).
  • the frame (2) can be constructed of a lightweight material, e.g., the frame (2) may comprise profiles made of a buoyant material selected from any of standard aluminum and its alloys, e.g., AICu, AlSi, AIMg, AlZn, AIMn, AITi, AINi, among others; plastic, e.g., polyvinyl chloride (PVC), polylactic acid (PLA), acrylonitrile butadiene styrene (ABS), polyethylene terephthalate (PET), polyethylene (PE), polypropylene (PP), expanded polystyrene (EPS), among others; wood or cork, among others with similar buoyant properties and not limited to the foregoing mentioned.
  • PVC polyvinyl chloride
  • PLA polylactic acid
  • ABS acrylonitrile buta
  • the system (1 ) may comprise floaters (3) that can help the frame (2) to increase and/or control the buoyancy of the system (1 ).
  • floaters (3) can be selected from any of foam floaters, air or gas filled floaters, and wooden floaters, or be made of any other element that provide to the frame (2) the capacity of floating.
  • Floaters (3) can be made of non-expensive materials that are globally available to ensure that the invention is cost-efficient, as well as to ensure scalability, replicability, and competitiveness.
  • floaters (3) are foam floaters that can be made of a material selected from one of polyethylene, polyurethane, or polystyrene. Because of the practical structure of the system (1 ), the same can be reused in a new growth cycle.
  • the floating rafting system (1 ) is placed within a container (8) filled with a liquid culturing medium (6) designed to allow the growth of a biomaterial (7) (see Figure 5) formed/secreted onto the substrate (5).
  • the container (8) according to the embodiment shown in Figure 1 has a size slightly greater than the size of the floating rafting system (1 ). In an embodiment, the container (8) has a size within a range of 1 to 100 mm greater than the size of the floating rafting system (1 ).
  • the container (8) can be made of any material compatible with the liquid culturing medium (6), so this does not affect the growing biomaterial process, and that is reusable, durable and cost-effective.
  • Some materials that can be used for the container (8) are any of the type of plastic, e.g., polyethylene terephthalate (PET), high-density polyethylene (HDPE), low-density polyethylene (LDPE), polyvinyl chloride (PVC), polystyrene (PS), polypropylene (PP), glass fiber reinforced polymers, stainless steel, among others.
  • PET polyethylene terephthalate
  • HDPE high-density polyethylene
  • LDPE low-density polyethylene
  • PVC polyvinyl chloride
  • PS polystyrene
  • PP polypropylene
  • glass fiber reinforced polymers stainless steel, among others.
  • the container is previously sterilized either chemically (e.g., using liquid solutions like hydrogen peroxide, or gas, like ethylene oxide, etc.)
  • the container (8) is not limited to a specific geometry. That is, the container (8) can be of rectangular, circular, triangular, trapezoidal or square shape, or any other suitable shape.
  • the floating rafting system (1 ) floats over the liquid culturing medium (6) allowing contact at least between the bottom surface of the substrate (5) with the liquid culturing medium (6).
  • FIGS. 1 and 2 show an embodiment of a floating rafting system (1 ) having rectangular geometry
  • FIG. 2 shows an embodiment of a floating rafting system (1 ) having rectangular geometry
  • floaters (3) are used and are joined beneath the frame (2) by using a commercial adhesive, which can be of the type of hotmelts, solvent- based, water-based, contact, polyacrylates, epoxy, polyurethanes, silicone adhesives among others.
  • floaters (3) might be placed on the lateral side of the frame (2) without affecting the gist of the invention.
  • floaters can be fastened to the frame mechanically by using any of screws, nuts and bolts, nails, or the like, avoiding the use of any glue.
  • the materials of the frame (2) and floaters (3) must be compatible to be easily attached, glued or somehow adhered together without the risk of detachment while the floating rafting system (1 ) is floating over the liquid culturing medium (6).
  • FIG 3 is an exploded view of the preferred embodiment of the floating rafting system (1 ) shown in Figures 1 and 2 in which the floating rafting system (1 ) is assembled with the substrate (5), frame (2), floaters (3) and fastening means (4).
  • the fastening means (4) can be tools or elements to properly hold the substrate (5) to the frame (2) of the floating rafting system (1 ).
  • the fastening means (4) are designed to be of a non-permanent joint, in order to easily replace the substrate (5) from the frame (2), without the substrate (5) suffering any damage, and without damaging the frame (2) and floaters (3), so the same can be reused in a new growth cycle.
  • the fastening means (4) can be selected from any of screws, nuts and bolts, clamps, hasp, anchors, rivets, pin, among others.
  • the fastening means (4) can be made of a hollow tubing or rigid material, so they do not affect the flotation of the frame (2) and floaters (3) over the liquid culturing medium (6).
  • fastening means (4) are, e.g., standard aluminum and its alloys, e.g., AICu, AlSi, AIMg, AlZn, AIMn, AITi, AINi, among other; stainless steel, and plastics, e.g., polyethylene terephthalate (PET), high-density polyethylene (HDPE), low-density polyethylene (LDPE), polyvinyl chloride (PVC), polystyrene (PS), polypropylene (PP), among others. Additionally, fastening means must be compatible with the frame in shape, composition and conformation.
  • standard aluminum and its alloys e.g., AICu, AlSi, AIMg, AlZn, AIMn, AITi, AINi, among other
  • stainless steel e.g., stainless steel, and plastics, e.g., polyethylene terephthalate (PET), high-density polyethylene (HDPE), low-density polyethylene (LDPE), polyviny
  • the fastening means (4) are designed to have the same geometry than the frame (2) in order to be aligned with the inner surface of the frame (2), so the substrate (5) is hold between the frame (2) and the fastening means (4) achieving the objective of having an accurate assembly and avoiding that substrate (5) moves or slips out.
  • the substrate (5) is arranged in the bottom surface of the frame (2) to allow the contact with the liquid culturing medium (6) (see Figure 1 ).
  • the substrate (5) is fastened to the frame (2) in such a manner that it is completely and evenly tensed, but without exerting too much pressure in the same to avoid damage.
  • floaters (3) which are elements used for increasing and/or controlling the buoyancy of the floating rafting system (1 ) can be arranged to the frame (2) using different configurations.
  • at least one floater (3) could be joined to each side of the frame (2).
  • Figure 4 shows a bottom view of the floating rafting system (1 ) showed in the embodiments of Figures 1 and 2, in which the substrate (5) is fixed to the frame (2) (see Figures 1 to 3) and the floaters (3) by the fastening means (4).
  • the fastening means (4) are disposed within the bottom surface of the frame (2).
  • the floaters (3) can be configured in a manner that a plurality of them can be attached to each side of the frame (2). It can be understood that the present invention can protect any other embodiment on how the floaters can be arranged, in order to keep the floating rafting system (1 ) floating over the liquid culturing medium (6).
  • the frame (2) can comprise fastening lanes (not shown) into which fastening means (4) can be inserted.
  • these fastening lanes can be configured as, but not limited to, a slit, opening, gap, or the like engraved on the frame (2). Those skilled in the art will understand that any variation on the fastening lanes will not affect the gist of the invention.
  • the fastening means (4) are standard vinyl spline (cord) which are inserted into fastening lanes (not shown) for assuring a good fastening of the substrate (5).
  • the substrate (5), which is fastened in the frame (2) can be a textile selected from a woven or a non-woven textile.
  • the woven and non-woven textile comprises any of plastic (e.g., polyester, nylon, acrylic, polylactic acid (PLA), polyhydroxybutyrate (PHB) polyamide); or any other materials comprising semi-synthetics such as rayon; mineral such as glass wool, mineral wool, rock wool; or viscose and natural or organic materials (e.g., cotton, bamboo, banana fiber, silk, wool, cellulose), without being limited to these materials only.
  • plastic e.g., polyester, nylon, acrylic, polylactic acid (PLA), polyhydroxybutyrate (PHB) polyamide
  • any other materials comprising semi-synthetics such as rayon; mineral such as glass wool, mineral wool, rock wool; or viscose and natural or organic materials (e.g., cotton, bamboo, banana fiber, silk, wool, cellulose), without being limited to these materials only.
  • the substrate becomes an embedded material which may include one or more materials.
  • the embedded material may be vegetable based, protein based, animal derived, plastic derived, organic, etc.
  • the substrate (5) can also be an electronic circuit, silicon circuit board, conductive thread, wire or similar to make smart textiles for wearable applications.
  • the floating rafting system (1 ) can comprise a supporting mesh (not shown) attached to the frame (2).
  • the supporting mesh can be made of any suitable rigid material such as plastic or metal.
  • the supporting mesh can have a sieve mesh size suitable not only to allow the liquid culturing medium to pass through it, but to superpose the substrate (5) evenly tensed thereon, when necessary.
  • the sieve mesh size can be of 10 mm or less.
  • the substrate (5) can be superposed on the supporting mesh without the need to be fastened directly to the frame (2).
  • a woven or non-woven textile or other suitable substrate can be placed evenly tensed over the supporting mesh without the need to be fastened directly to the floating rafting system (1 ) because the tension of the supporting mesh is transferred to the textile substrate.
  • the substrate (5) can consist of loose fibers widespread over the supporting mesh.
  • the loose fibers can be selected from plastic (e.g., polyester, nylon, acrylic, polylactic acid (PLA), polyhydroxybutyrate (PHB) polyamide); or any other materials comprising semi-synthetics such as rayon; mineral such as glass wool, mineral wool, rock wool; or viscose and natural or organic materials (e.g., cotton, bamboo, banana fiber, silk, wool, cellulose), Kevlar fibers, carbon fibers, lignocellulose fibers, among others, and combinations thereof without being limited to these materials only.
  • plastic e.g., polyester, nylon, acrylic, polylactic acid (PLA), polyhydroxybutyrate (PHB) polyamide
  • PHA polylactic acid
  • PHB polyhydroxybutyrate
  • a method to grow a biomaterial (7) directly onto a substrate (5) comprises the steps of: providing a floating rafting system (1 ) according to the current invention, placing the floating rafting system (1 ) into a liquid culturing medium (6), letting the biomaterial (7) to incubate and grow, removing the floating rafting system (1 ) from the liquid culturing medium (6) and separating the substrate (5) comprising the biomaterial (7) grown from the floating rafting system (1 ).
  • the substrate (5) is not totally submerged into the liquid culturing medium (6) allowing the biomaterial (7) to grow on the opposite side contacting said liquid culturing medium (6).
  • the substrate (5) is fully submerged into the liquid culturing medium (6), but wherein the floating rafting system (1 ) is still floating over the liquid culturing medium.
  • the liquid culturing medium (6) is enriched with cells.
  • a culturing medium is a set of nutrients, growth factors and other components (e.g., agar, extracts (from some animals or vegetables), peptones, bodily fluids, buffer systems, pH indicators, reducing agents and selective agents), that create the necessary conditions for development of cells such as mammalian cells, fish cells, amphibian cells, insect cells, plant cells, bacterial cells and fungal cells, including yeast cells and mold cells.
  • agar e.g., agar, extracts (from some animals or vegetables), peptones, bodily fluids, buffer systems, pH indicators, reducing agents and selective agents
  • cells such as mammalian cells, fish cells, amphibian cells, insect cells, plant cells, bacterial cells and fungal cells, including yeast cells and mold cells.
  • the metabolic diversity of the same is so great that the variety of culturing medium is enormous, since there is no universal crop suitable for all of them, not even referring to bacteria exclusively.
  • the liquid culturing medium is a bacterial culturing medium which comprises dried food waste, dried black tea, and water.
  • Table 1 below is an illustrative example of the constitution of the culturing medium according to the invention:
  • the biomaterial can grow onto the substrate (5) until the liquid culturing medium (6) has no nutrients allowing the biomaterial to grow. Additionally, it should be taken into account that certain quantity of the liquid culturing medium (6) might evaporate during the growth process. It is possible to add liquid culturing medium (6) to the container (8), for example, every day during the incubation process, in order to always have an adequate level of liquid culturing medium (6) in the container (8). Once the biomaterial has grown onto the substrate (5), the floating rafting system (1 ) is removed from the culturing medium (6).
  • the floating rafting system is flipped 180° and is left floating into the liquid culturing medium for another growth period, so there is an even growth of biomaterial throughout both sides of the substrate.
  • the substrate (5) comprising the biomaterial (7) is separated from the floating rafting system (1 ).
  • the obtained product (substrate and biomaterial) is post-processed for giving the desired technical properties according to the final use.
  • Said post-processing may include a tanning process.
  • the final product can be used for the manufacture of daily items like shoes, clothes, handbags, wallets, or the similar.
  • the biomaterial (7) generated is based on bacterial cellulose (BC).
  • BC is an insoluble extracellular polymer, which is produced by various species of bacteria, such as those belonging to the genera Acetobacter, Rhizobium, Agrobacterium and Sarcina.
  • the genus Acetobacter encompasses a group of bacteria with the ability to oxidize sugars and ethanol generating acetic acid.
  • Among the most effective cellulose include A. xylinum, A. hansenii, and A. pasteurianus. From among them A. xylinum has been taken as a model organism for the studies basic and applied on BC, due to its ability to produce, starting from various carbon sources, high levels of the polymer as primary product of its metabolism.
  • BC is a hydrogel composed from a nano assembled, polysaccharide matrix, of pure cellulose microfibrils, e.g., microbial cellulose, and nano cellulose.
  • BC has distinctive and superior properties compared to vegetal cellulose, in terms of purity, resistance, malleability and capacity of water retention, among others, making this material acceptable for applications in fields such as biotechnology, microbiology, and material science.
  • the main bacterium considered for generating the adequate growing conditions is the Acetobacter xylinum (or Komagataeibacter xylinus) from any of the following group: Komagataeibacter sp, Gluconacetobacter sp, Acetobacter sp, Rhizobium sp, Sarcina sp, Pseudomonas sp, Achromobacter sp, Alcaligenes sp, Aerobacter sp, and Azotobacter sp; which is a species of bacteria best known for its ability to produce cellulose, specifically bacterial cellulose. K.
  • xylinus is a member of the acetic acid bacteria, a group of Gram-negative aerobic bacteria that produce acetic acid during fermentation. K. xylinus is unusual among the group in also producing cellulose. Bacterial cellulose (also sometimes known as nanocellulose) is involved in the formation of biofilms. It is chemically identical to plant cellulose but with distinct physical structure and properties.
  • the present invention refers to a method to grow a BC directly onto a substrate (5).
  • the obtained biomaterial (7) has mechanical and physical properties adequate for the manufacture of the daily items.
  • the following table 2 in an illustrative manner, shows an example of the mechanical properties that the biomaterial (7) presents.
  • the tensile strength was determined by using a Universal Machine Chatillon LR 10K 1200 Ibf and according to ASTM D638:
  • FIG 5 is a perspective view of an embodiment of the floating rafting system (1 ) showing the biomaterial (7) grown onto a substrate (not visible) within the floating rafting system (1 ).
  • This is an exemplary drawing showing a layer of biomaterial (7) that has grown evenly after the floating rafting system (1 ) was left in a liquid culturing medium (6) (please see Figure 1 ).
  • This layer of biomaterial (7) covers uniformly all the substrate disposed between the frame (2) and the fastening means (4).
  • One advantage of using the floating rafting system (1 ) of the present invention is that the same can be used as many times as desired replacing the substrate (5) having the biomaterial (7) for a new one and using a new liquid culturing medium (6).
  • the substrate (5) having the biomaterial (7) presents good bonding and fiber intertwining, feature which is critical for avoiding a lamination step and for giving to the final items the technical properties, e.g. high resistance, tensile strength, flexibility and others.
  • the biomaterial (7) is extracted from the floating rafting system (1 ), it could be subjected to a post-processing step in order to produce a commercial product.
  • the most preferable post-processing process is tanning, but other or further post-processing steps can also be performed.
  • the present specification and drawings of embodiments, in accordance with the gist of the present invention are merely illustrative of the principles of the invention. Therefore, it should be understood that variations and/or modifications can be carried out by those skilled in the art without implying that such modifications depart from the spirit and scope of the present invention. Accordingly, the invention is not limited except as by the appended claims.

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Abstract

La présente invention concerne un procédé de croissance d'un biomatériau (7) directement sur un substrat (5) à l'aide d'un système de chevron flottant (1) sur un milieu de culture liquide (6) pour incuber ledit biomatériau (7) empêchant le besoin de stratifier le biomatériau (7) sur le substrat (5) ; le biomatériau (7) obtenu avec ledit procédé, et un système de chevron flottant (1) utilisé dans celui-ci. De même, la présente invention concerne un système de chevron flottant (1) destiné à être utilisé dans le procédé mentionné ci-dessus, le système de chevron flottant (1) comprenant un cadre (2) et un substrat (5).
PCT/IB2022/054415 2022-05-12 2022-05-12 Procédé de croissance d'un biomatériau sur un substrat à l'aide d'un système de chevron flottant WO2023218226A1 (fr)

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