[go: up one dir, main page]

WO2025048697A1 - Procédé de culture d'aliments et de boissons dans un bioréacteur - Google Patents

Procédé de culture d'aliments et de boissons dans un bioréacteur Download PDF

Info

Publication number
WO2025048697A1
WO2025048697A1 PCT/SE2024/050736 SE2024050736W WO2025048697A1 WO 2025048697 A1 WO2025048697 A1 WO 2025048697A1 SE 2024050736 W SE2024050736 W SE 2024050736W WO 2025048697 A1 WO2025048697 A1 WO 2025048697A1
Authority
WO
WIPO (PCT)
Prior art keywords
media
culture tank
bioreactor
sterilized
cells
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/SE2024/050736
Other languages
English (en)
Inventor
Marie GIBBONS
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Re meat AB
Original Assignee
Re meat AB
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Re meat AB filed Critical Re meat AB
Publication of WO2025048697A1 publication Critical patent/WO2025048697A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
    • C12M29/26—Conditioning fluids entering or exiting the reaction vessel
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0011—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using physical methods
    • A61L2/0029—Radiation
    • A61L2/0047—Ultraviolet radiation
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M27/00—Means for mixing, agitating or circulating fluids in the vessel
    • C12M27/02—Stirrer or mobile mixing elements
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M31/00—Means for providing, directing, scattering or concentrating light
    • C12M31/10—Means for providing, directing, scattering or concentrating light by light emitting elements located inside the reactor, e.g. LED or OLED
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M37/00—Means for sterilizing, maintaining sterile conditions or avoiding chemical or biological contamination
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2202/00—Aspects relating to methods or apparatus for disinfecting or sterilising materials or objects
    • A61L2202/10—Apparatus features
    • A61L2202/11—Apparatus for generating biocidal substances, e.g. vaporisers, UV lamps

Definitions

  • the present disclosure relates to the field of methods for cultivating food and beverages in a bioreactor and to a bioreactor for use in the method.
  • Cultivated meat is an emerging industry that aims to provide consumers with meat that is produced through animal cell cultivation, rather than traditional animal agriculture.
  • the production of cultivated meat involves growing animal cells in a bioreactor, which is a vessel designed to support the growth and differentiation of cells in a controlled environment.
  • one object of the present invention is to provide a method for cultivating food and beverages in a bioreactor, which increases the efficacy, lowers the costs and provides for a final food or beverage product that both meets the high regulatory demands for such products and improves consumer acceptance.
  • a method for cultivating food and beverages in a bioreactor comprises the steps of; a) preparing a media, the media being any one of; a basal media, a growth media or a differentiation media; b) sterilizing the media prepared in step a); c) sterilizing a culture tank and one or more ports of a bioreactor by using one or more UV light sources; d) providing the media into the culture tank of the bioreactor via the one or more port(s), wherein, step d) may alternatively be carried out prior to step b) followed by a simultaneous sterilization of the culture tank and the media in step c) or according to a further alternative, step d) is carried out subsequent to step c) and after these are performed, the media is sterilized according to step b) in the culture tank by using the one or more UV light sources; e) adding meat cells or non-photosynthetic cells to the sterilized media, either prior to introducing the sterilized media into the culture
  • the present invention relates to a method for the cultivating of animal cells in the production of cultivated meat.
  • the bioreactor for use in the method utilizes ultraviolet (UV) radiation as a cost-effective and rapid sterilization method, eliminating the need for traditional steam-in- place processes.
  • UV radiation optionally in combination with other food-safe sterilization methods, ensures a high level of germ eradication, facilitating a safe and efficient bioreactor system.
  • the bioreactor can be constructed using alternative materials other than stainless steel or single use plastic, which is the industry standard. Using materials such as multi-use plastic and glass reduces manufacturing costs and enables larger scalability.
  • the method as disclosed herein may comprise a step of sterilizing the cell product by using a UV light source.
  • the step of sterilizing the cell product by using a UV light source inactivates 90% or more of the cells in the cell product, based on the numbers of the cells, alternatively or additionally wherein the step of sterilizing the cell product by using a UV light source is carried out such that the level of contaminants in the cell product is 0. 1 % or lower, or 0.01% or lower, based on the number of the contaminants compared to a reference sample.
  • the number of inactivated cells and the level of contaminants in the cell product is measured by a “proliferation assay” and is compared to a reference sample.
  • Standardization refers to the act of inhibiting cells and other contaminants from replicating. This can include but is not limited to animal cells, plant cells, bacteria, yeast, viruses, and spores.
  • cultiva is meant a product containing cells isolated from an animal and subsequently grown outside of the animal for the purpose of consumption.
  • CIP/cold sterilization involves the use of acids, bases, and other food-safe reagents such as peracetic acid and hydrogen peroxide to achieve thorough cleaning and preliminary microbial reduction. These reagents are carefully selected for their efficacy and safety in food processing environments.
  • the CIP/cold sterilization process primarily serves to remove organic and inorganic residues, reduce microbial load, and prepare surfaces for subsequent UV sterilization.
  • the mixing step a) may be carried out in separate equipment such as a mixing tank or alternatively/ additionally inside the culture tank of the bioreactor.
  • the mixing is preferably carried out until all nutrients, growth and/or differentiation factors are solubilized.
  • the growth media or the differentiation media may further comprise a buffer.
  • the culture tank, and the port(s) of the bioreactor may be decontaminated, such as by using an acid, a base, gas, and/or ethanol.
  • the one or more port(s) of the bioreactor may for example be entry port(s), exit port(s), sampling port(s), a port for a sensor or probe to enter or exit the tank or a port allowing gas exchange, i.e., the port as defined herein is an opening of the bioreactor allowing access to the culture tank.
  • Step d) of introducing the media into the culture tank may be carried out subsequent to step c), alternatively step d) may be carried out prior to the step c) of sterilizing the culture tank of the bioreactor, such that the culture tank and the media is sterilized simultaneously by the UV light source(s), i.e. such that step b) and c) are carried out at the same time and preferably by UV light sources being integrated within the culture tank.
  • the UV light source(s) i.e. such that step b) and c) are carried out at the same time and preferably by UV light sources being integrated within the culture tank.
  • Step c) of sterilizing the culture tank, if the media is sterilized separately, may be carried out prior to, simultaneously with or after steps a) and b).
  • the media should be sterilized prior to adding the meat cells or non-photosynthetic cells to the media.
  • One option is to add the meat cells or non-photosynthetic cells to the sterilized media prior to adding the media comprising the cells into the sterilized culture tank, i.e.
  • one alternative is to sterilize the media and the culture tank separately and then add the sterilized media into the sterilized culture tank and in a subsequent step to add the meat cells or non-photosynthetic cells to the media in the culture tank
  • a still further alternative is to add the media into the culture tank and simultaneously sterilize the media and the culture tank, as described above, and in a subsequent step add the meat cells or non-photosynthetic cells to the sterilized media in the culture tank and in a still further step the meat cells or non-photosynthetic cells are added to a sterilized culture tank, i.e. sterilized culture tank via a sterilized port, and in a subsequent step, sterilized media is added to the sterilized culture tank.
  • the media prepared in step a) may be a basal media, a growth media or a differentiation media, the basal media being a base for preparing either one of a growth media or a differentiation media and comprises at least water and nutrients.
  • the adding of either growth factors to prepare a growth media or differentiation factors to prepare a differentiation media may be carried out directly after step b) of sterilizing the media, after step d) of providing the media into the culture tank and also prior to, or after adding the meat cells or non-photosynthetic cells to the sterilized media.
  • Steps f) and g) are optional steps and depend on the cell type utilized. Certain cell types require growth factors and certain cell types naturally do not require cell factors to grow or have been genetically or epigenetically modified to eliminate the need for growth factors.
  • Pipes/tubings included in, or connected to, the bioreactor may also be sterilized, such as by using UV radiation, heat, peracetic acid (PAA), iodophor, phosphoric acid, dodecylbenzenesulfonic acid, chlorine, chlorine dioxide, quaternary ammonium compounds, isopropanol, ethanol, Sodium hydroxide, gas, ozone, or other food-safe sterilization methods.
  • PAA peracetic acid
  • iodophor iodophor
  • phosphoric acid phosphoric acid
  • dodecylbenzenesulfonic acid iodophor
  • chlorine chlorine dioxide
  • quaternary ammonium compounds isopropanol
  • ethanol ethanol
  • Sodium hydroxide gas
  • gas ozone
  • other food-safe sterilization methods such as by using UV radiation, heat, peracetic acid (PAA), iodophor, phosphoric acid, dodecylbenzenesul
  • the bioreactor used in the method as disclosed herein may be completely or partially made of plastic material(s).
  • the culture tank, the port(s) and/or pipes/tubings may be made entirely, or at least partially, by plastic material(s).
  • 50% or more of the bioreactor is made of plastic material(s).
  • the plastic material may for example be a polyolefin plastic material, a polycarbonate, polyvinyl chloride, polymethyl methacrylate, polylactic acid and/or polyethylene terephthalate material.
  • the meat cells or non-photosynthetic cells have been immortalized and/or have a doubling time of less than 48h and/or are capable of growing in single cell suspension, and/or can grow in the presence of inhibitory metabolites, and/or are able to grow and/or differentiate without growth and/or differentiation factors.
  • the benefit of using UV-sterilization of such meat cells or non-photosynthetic cell is that the UV light prevents the cell product from further post-cultivation replication, which has otherwise been a consumer concern with the present type of products.
  • the time of sterilizing the culture tank and the one or more ports of the bioreactor and/or the cell product is within the range of from 1 sec to 72h.
  • the time of sterilizing the culture tank and the one or more ports of the bioreactor and/or the cell product is within the range of from 10 minutes to 24 hours, 15 minutes to 5 hours, or preferably 30 minutes to 3 hours.
  • ranges may vary depending on different material sensitivities to UV radiation, different areas of the reactor can benefit from being irradiated for different times and with intensities.
  • the supply of nutrients and growth and/or differentiation factors may be ceased prior to sterilizing the cell product.
  • the cells are stopped from growing. This may be done once the limits of the reactor size and viscosity of the media are reached. This may also trigger the cells to differentiate into, for example, muscle fibers. A further advantage is that it may lead to cell death, which might be important for regulations and/or consumer acceptance.
  • the UV spectrum upon sterilization is within the range of from 100 and 280 nm, preferably within the range of from 240 and 265 nm. This is particularly suitable for sterilizing the culture tank and the one or more ports of the bioreactor for cultivating meat or non-photosynthetic cells.
  • the one or more port(s) of the bioreactor has/have been sterilized with UV light prior to introducing the media or the cells into the port(s). It has been found by the present inventor that the port(s) of the bioreactor may be a source of contamination for the media or the cells entering the culture tank since the port(s) is/are a semi-external environment and sterilizing the port(s) of the bioreactor with UV light prior to introducing the media or cells into the port(s) has been found to significantly reduce the risk of contamination.
  • the bioreactor comprises a sensor system and the method comprises a step of monitoring the effectiveness of the UV light exposure, such as measuring the wavelength emitted by the UV light source.
  • the sensor system may for example by arranged in the culture tank of the bioreactor. This will ensure a continuous and high level of germ eradication and sterilization through-out large-scale production.
  • a sensor system for use in the present method may furthermore monitor, and optionally selfregulate, any one or all of the following; the temperature, pH, the level of nutrients, metabolites, oxygen, air and/or carbon dioxide. This may enhance the conditions for the cultivation process and thus speed up or improve the quality of the final cell product.
  • the sterilization in step b) is performed via filtration and the filter has a pore size within the range of from 0.1 to 0.5 pm, preferably within the range of from 0.15 to 0.3 pm. This range allows for critical nutrients such as amino acids, vitamins, minerals, lipids, carbohydrate sources, and growth and/or differentiation factors to pass through the filter, while inhibiting passage of contaminants such as yeast and bacteria which are too large to fit through the filter membranes.
  • the sterilization of the media may be carried out by heat, gas, ozone or UV radiation.
  • the culture tank of the bioreactor comprises two or more UV light sources.
  • the UV light source(s) arranged in the culture tank is/are ring-shaped UV light sources.
  • the ring-shaped UV light sources may be continuous UV light sources, such as a continuous circumferential light source.
  • ring-shaped herein includes circular, oval, rectangular and other, for example, rhomboidal ring-shapes.
  • the ring-shape may be a continuous or a discontinuous ring-shape, with “discontinuous ring-shape” herein meaning separate UV light sources forming the outline of a ring-shape.
  • the culture tank of the bioreactor has a top section and a bottom section, as seen in a hight direction, and wherein each one of the top section and the bottom section comprises a respective ring-shaped UV light source.
  • the culture tank has an inner diameter, as measured between the opposing inner walls of the culture tank and wherein an outer diameter of the ring-shaped UV light sources, is within the range of from 20% to 95% of the inner diameter of the culture tank, optionally within the range of from 25% to 85% of the inner diameter of the culture tank, preferably within the range of from 40% to 75% of the inner diameter of the culture tank .
  • the width of the UV-light source, or the UV-light bulb is the same or greater than the distance between the inner wall of the culture tank and the UV-light source/UV-light bulb.
  • the cell lines introduced in the media are animal cells and the final food product is cultivated meat.
  • growth media When growth media is used, this is a liquid medium used to promote and support cell proliferation.
  • Growth media generally comprises water, nutrients and growth factors.
  • An example would a be a proliferation media formulation that promotes cells to grow from 100,000 cells/mL of media to 10,000,000 cells/mL of media over 5 days.
  • Differentiation media is a liquid medium used to promote and support cell differentiation and maturation.
  • Differentiation media generally comprises water, nutrients and differentiation factors.
  • An example would be a differentiation media formulation that promotes myoblasts to align and fuse together to form myofibers.
  • basic media is meant a media comprising water and nutrients and which forms a basis for preparing either a growth or a differentiation media.
  • culture tank or “culture tank” is meant either both the tank itself and any therein contained equipment and/or components, including but not limited to i.e. agitators, spargers, baffles or probes, or only the tank itself, as used when describing i.e. dimensions of the tank.
  • This can also include but is not limited to membranes, tubes, piping, fittings, or any other equipment that may be connected to the culture tank or bioreactor, and/or may come in contact with the media, cells, and/or product.
  • the present disclosure relates to a bioreactor for use in a method according to the first aspect, wherein the bioreactor comprises one or more ports and a culture tank, wherein the culture tank comprises an upper portion and a bottom portion, as seen in a height direction (H), wherein an upper ring-shaped UV light source is arranged in the upper portion of the culture tank and a lower ring-shaped UV light source is arranged in the lower portion of the culture tank.
  • H height direction
  • the bioreactor may be completely or partially be made of a plastic material.
  • the culture tank, the port(s) and/or the pipes may be made entirely or at least partially by plastic material.
  • 50% or more of the bioreactor is made of plastic material.
  • the plastic material may for example be a polyolefin plastic material, a polycarbonate, polyvinyl chloride, polymethyl methacrylate, polylactic acid and/or polyethylene terephthalate plastic material.
  • ring-shaped herein is intended both circular, oval, rectangular or other, for example, rhomboidal ring-shape.
  • the ring-shape may be a continuous or a discontinuous ring-shape, with “discontinuous ring-shape” herein meaning separate UV light sources forming the outline of a ring-shape.
  • the culture tank has an inner diameter, as measured between the opposing inner walls of the culture tank and wherein an outer diameter of the ring-shaped UV light sources, is within the range of from 25% to 95% of the inner diameter of the culture tank, preferably within the range of from 25% to 85% or 45% to 75%. It has been found by the present inventors that this improves the exposure and optimize the efficiency in terms of time and energy consumption.
  • the width of the UV-light source is the same or greater than the distance between the inner wall of the culture tank and the UV-light source.
  • the culture tank comprises one or more UV reflectors.
  • the UV reflectors are arranged at or on an inner side wall of the culture tank.
  • the culture tank comprises a circumferential UV reflector section arranged continuously or discontinuously around the circumference of the inner side wall of the culture tank.
  • the one or more UV reflectors may alternatively or additionally be arranged at or on an inner top or bottom wall of the culture tank.
  • the reflectors are arranged in the center of the culture tank.
  • the reflectors are attached to an agitator or other component within the culture tank or reactor.
  • Fig. 1 is a schematic illustration of the method according to the present disclosure
  • Fig. 2 illustrates an exemplary embodiment of a bioreactor for use in a method according to the present disclosure
  • Fig. 3a-d illustrate different exemplary embodiments of bioreactors for use in the method according to the present disclosure.
  • FIG. 1 is a schematic illustration of the method according to the present disclosure wherein the method comprises the steps of; a) preparing a media, the media being any one of; a basal media, a growth media or a differentiation media; b) sterilizing the media prepared in step a); c) sterilizing a culture tank and one or more ports of a bioreactor by using one or more UV light sources; d) providing the media into the culture tank of the bioreactor via the one or more port(s), wherein, step d) may alternatively be carried out prior to step b) followed by a simultaneous sterilization of the culture tank and the media in step c) or according to a further alternative, step d) is carried out subsequent to step c) and after these are performed, the media is sterilized according to step b) in the culture tank by using the one or more UV light sources; e) adding meat cells or non-photosynthetic cells to the sterilized media, either prior to introducing the sterilized media into the culture tank in step d
  • FIG. 2 illustrates an exemplary embodiment of a bioreactor 1 for use in a method according to the present disclosure.
  • the bioreactor 1 comprises a first and a second port 2,3 and a culture tank 4.
  • the culture tank 4 comprises an upper portion 5 and a bottom portion 6, the culture tank furthermore comprises a side wall 7, extending between a bottom 6a and a top 5a of the culture tank 4, as seen in a height direction H of the bioreactor 1.
  • the culture tank 4 furthermore comprises an inner side 8 and an outer side 9.
  • the bioreactor 1 in this Figure 2 is a stirred tank bioreactor comprising an agitation system 10 including an agitator 11 and a motor 12. This is however an optional feature, and the bioreactor may alternatively be of any bioreactor configuration, including but not limited to an air lift reactor, a bubble column reactor, a hollow filter membrane reactor or a non-agitating reactor.
  • a heating/cooling jacket 13 is enclosing the bioreactor 1 on the outer side 9 of the bioreactor 1 to provide a stable temperature suitable for cultivation according to the present method.
  • a suitable temperature may for example be within the range of from 30 to 45°C.
  • the ports 2,3 are connected to a respective piping/tubing 14,15 for feeding the medium (not shown) into the culture tank 4, such as from a separate mixing tank.
  • the respective piping/tubing 14,15 may be fixed or detachably attached to the respective port 2,3.
  • the culture tank 4 is provided with an upper UV light source 17 and a bottom UV light source 18.
  • the UV light sources 17,18 are ring-shaped UV light sources.
  • the upper and/or bottom UV light source 17,18 may be an integral part of the culture tank 4, such as being arranged flush, or essentially flush, with the respective top and the bottom 5a, 6a of the culture tank 4 or with the side wall 7.
  • the culture tank 4 has an inner diameter d4, as measured between opposing inner walls 7a, 7b of the culture tank 4 and wherein the respective outer diameter dn/dis of the ring-shaped UV light sources 17,18, is within the range of from 20% to 95%, optionally within the range of from 25% to 85%, preferably within the range of from 40% to 75%, of the inner diameter d4 of the culture tank 4. This has been found advantageous for providing a rapid and uniform sterilization within the culture tank 4.
  • One or both of the UV-light source(s) 17,18 may have a width wn/wis, as measured in a direction perpendicular to the height direction H of the bioreactor 1, which may be the same or greater than a distance ds between the inner side 8 of the culture tank 4 and the UV-light source 17,18.
  • a sterile tube or pipe 21 may be connected to an exit port 22 of the culture tank 4 and a decanter centrifuge may be connected to the culture tank 4 via the sterile tube or pipe 21.
  • the bioreactor 1 in Fig. 3b comprises a single UV light source 17 extending from a central portion of the top 5 a of the culture tank 4.
  • the bioreactor in Fig. 3c is provided with an upper central UV light source 17 arranged over a shaft of the agitator 11 of the bioreactor 1.
  • the bioreactor 3c furthermore comprises a bottom UV light source 18 extending from the bottom 6a of the bioreactor 1.
  • Clean-in-place was performed on an empty 500-liter media mixing tank.
  • the mixing tank was closed and cold water was added for 3-5 minutes.
  • the port at the bottom of the tank was used to allow draining.
  • Wash #1 was performed using a detergent for 5-15 minutes, and was allowed to drain, and then rinsed again with cold water for 3 minutes.
  • the mixing tank was allowed to drain and wash #2 was performed using dilute acid (0.5-1%) for 5-15 minutes.
  • the mixing tank was again allowed to drain, and the final rinse was performed using de-ionized water for 3 minutes.
  • the media mixing tank was allowed to drain completely before opening the tank and dry modified E8 media powder was added to the media mixing tank, followed by 400 liters of deionized water.
  • the media was mixed using a propeller agitator at 100RPM for 20 minutes. It was confirmed that the media powder was completely solubilized by collecting and observing a sample. Once the dry powder was solubilized, 4.28 grams of transferrin, 0.04 grams of fibroblast growth factor 2, 0.004 grams of insulin-like growth factor 1, and 0.0008 grams of transforming growth factor pi were added to the media.
  • Clean-in-place was performed on an empty 500-liter bioreactor.
  • the bioreactor was closed, and cold water was added for 3-5 minutes.
  • the port at the bottom of the bioreactor tank was used to allow draining.
  • Wash #1 was performed using a detergent for 5-15 minutes, and was allowed to drain, and then rinsed again with cold water for 3 minutes.
  • the bioreactor tank was allowed to drain and wash #2 was performed using dilute acid (0.5-1%) for 5-15 minutes.
  • the bioreactor tank was again allowed to drain, and the final rinse was performed using de-ionized water for 3 minutes.
  • the bioreactor tank was allowed to drain completely before sterilizing the tank with UV radiation.
  • UV sterilization was subsequently performed on the empty bioreactor by turning on the two ring-shaped UV-C bulbs located at the top and bottom of the tank.
  • the reactor was treated for 30 minutes at a wavelength of 254nm.
  • a sterilized hose containing a 0.22um polyethersulfone (PES) membrane filter was connected from the port at the bottom of the media mixing tank to the port at the top of the bioreactor.
  • PES polyethersulfone
  • suspension-adapted bovine myoblast cells generated from a seed train were aseptically added to the reactor at a density of 100,000 cells/mL through a port at the top of the reactor.
  • Cells were agitated for 5 days under the following culture conditions: 37°C temperature, 95% air and 5% carbon dioxide gas, 100RPM, and 7.4pH. The pH was maintained at 7.4 via pH probe monitoring and automatic addition of sodium bicarbonate.
  • a sterile tube and a decanter centrifuge were connected to the port at the bottom of the reactor and the sterilized media and cells were pumped into the centrifuge at a flow rate per bowl size of 0.75/min and spun at 1000G.
  • the solid biomass was subsequently collected from the centrifuge and a wash step was performed by re-suspending the cells in four-fold w/v of phosphate buffered saline. A further separation step was then carried out via centrifugation.
  • the resulting biomass should yield 4,000g of cultivated beef.
  • the cultivated beef was then fed into a twin-screw high moisture extruder along with the following seasonings to produce a minced cultivated beef product: salt, pepper, brown sugar, onion, garlic, cumin, cayenne pepper, paprika, chipotle powder, and vegan natural beef flavor.
  • the minced cultivated beef was sealed and stored for further preparations, analysis, packaging, and distribution.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Organic Chemistry (AREA)
  • Biomedical Technology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Genetics & Genomics (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Sustainable Development (AREA)
  • Molecular Biology (AREA)
  • Medicinal Chemistry (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Epidemiology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

La présente divulgation concerne un procédé de culture d'aliments et de boissons dans un bioréacteur (1), le procédé comprenant les étapes suivantes : - préparation d'un milieu, le milieu étant l'un quelconque des milieux suivants : milieu de base, milieu de croissance ou milieu de différenciation ; - stérilisation du milieu préparé à l'étape a) ; - stérilisation d'un réservoir de culture (4) et d'un ou plusieurs orifices (2,3,22) d'un bioréacteur (1) par l'utilisation d'une ou plusieurs sources de lumière UV (17,18) ; - mise en place du milieu dans le réservoir de culture (4) du bioréacteur (1) par l'intermédiaire d'un ou plusieurs orifices (2,3) ; - ajout de cellules de viande ou de cellules non photosynthétiques au milieu stérilisé, soit avant l'introduction du milieu stérilisé dans le réservoir de culture (4), soit en combinant les cellules de viande ou les cellules non photosynthétiques et le milieu stérilisé dans le réservoir de culture stérilisé (4), ou dans un équipement en relation avec le réservoir de culture stérilisé ; - ajout éventuel de facteurs de croissance ou de facteurs de différenciation au milieu, si le milieu est un milieu de base pour la préparation d'un milieu de croissance ou d'un milieu de différenciation ; - laisser les cellules de viande ou les cellules non photosynthétiques présentes dans le milieu stérilisé proliférer ou se différencier dans la cuve de culture (4) pour fournir un produit cellulaire ; - récolte et préparation d'un produit alimentaire et/ou d'une boisson précurseurs ou finaux à partir du produit cellulaire.
PCT/SE2024/050736 2023-08-25 2024-08-19 Procédé de culture d'aliments et de boissons dans un bioréacteur Pending WO2025048697A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
SE2350997 2023-08-25
SE2350997-9 2023-08-25

Publications (1)

Publication Number Publication Date
WO2025048697A1 true WO2025048697A1 (fr) 2025-03-06

Family

ID=94819920

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/SE2024/050736 Pending WO2025048697A1 (fr) 2023-08-25 2024-08-19 Procédé de culture d'aliments et de boissons dans un bioréacteur

Country Status (1)

Country Link
WO (1) WO2025048697A1 (fr)

Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008157247A1 (fr) * 2007-06-15 2008-12-24 Amgen Inc. Procédés de traitement de milieux de cultures cellulaires à utiliser dans un bioréacteur
WO2011153288A1 (fr) * 2010-06-01 2011-12-08 Alexander Farren Stérilisation uv de récipients
WO2018227016A1 (fr) * 2017-06-07 2018-12-13 Wild Type, Inc. Production de chair ex vivo
CN109468229A (zh) * 2018-12-05 2019-03-15 山西云度知识产权服务有限公司 一种用于生物医药生产的细胞培养装置
WO2020222239A1 (fr) * 2019-05-02 2020-11-05 Aleph Farms Ltd. Systèmes et procédés de culture pour la production à grande échelle d'aliments cultivés
EP3757201A1 (fr) * 2019-06-27 2020-12-30 Phytolinc UG Bioréacteur et procédé de traitement d'au moins un fluide et/ou de culture d'organismes phototrophes
US20220056394A1 (en) * 2020-08-18 2022-02-24 Upside Foods, Inc. Systems, devices, and methods for sterilizing bioreactors and culture media
WO2023012523A1 (fr) * 2019-12-02 2023-02-09 Avant Meats Company Limited Système de production de viandes, de tissus et de produits associés cultivés à partir de cellules
WO2023196089A1 (fr) * 2022-04-04 2023-10-12 Arcology Inc. Dba Biosphere Bioréacteurs configurés pour la stérilisation par uv, et procédés d'utilisation de la stérilisation par uv dans des bioprocessus

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008157247A1 (fr) * 2007-06-15 2008-12-24 Amgen Inc. Procédés de traitement de milieux de cultures cellulaires à utiliser dans un bioréacteur
WO2011153288A1 (fr) * 2010-06-01 2011-12-08 Alexander Farren Stérilisation uv de récipients
WO2018227016A1 (fr) * 2017-06-07 2018-12-13 Wild Type, Inc. Production de chair ex vivo
CN109468229A (zh) * 2018-12-05 2019-03-15 山西云度知识产权服务有限公司 一种用于生物医药生产的细胞培养装置
WO2020222239A1 (fr) * 2019-05-02 2020-11-05 Aleph Farms Ltd. Systèmes et procédés de culture pour la production à grande échelle d'aliments cultivés
EP3757201A1 (fr) * 2019-06-27 2020-12-30 Phytolinc UG Bioréacteur et procédé de traitement d'au moins un fluide et/ou de culture d'organismes phototrophes
WO2023012523A1 (fr) * 2019-12-02 2023-02-09 Avant Meats Company Limited Système de production de viandes, de tissus et de produits associés cultivés à partir de cellules
US20220056394A1 (en) * 2020-08-18 2022-02-24 Upside Foods, Inc. Systems, devices, and methods for sterilizing bioreactors and culture media
WO2023196089A1 (fr) * 2022-04-04 2023-10-12 Arcology Inc. Dba Biosphere Bioréacteurs configurés pour la stérilisation par uv, et procédés d'utilisation de la stérilisation par uv dans des bioprocessus

Similar Documents

Publication Publication Date Title
AU2015372409B2 (en) Method and device for treating foods and/or containers by means of a process liquid
JPS60500405A (ja) 次亜塩素酸塩の水溶液を用いて物品を消毒する装置
CN109907109A (zh) 一种基于正渗透原理的乳制品制备方法
KR102130662B1 (ko) 버섯종균의 대량생산이 가능한 배양장치
WO2025048697A1 (fr) Procédé de culture d'aliments et de boissons dans un bioréacteur
JP4567737B2 (ja) 酸乳製品の製造法、そのための乳の処理方法、酸乳製造ライン、該ラインにおける乳の処理装置
CN104353090B (zh) 一种用于无土栽培营养液的消毒净化系统及其净化方法
CN213924873U (zh) 一种规模化生产悬浮细胞的反应器
CN120505197A (zh) 一种智能化car-t细胞培育系统
KR101135052B1 (ko) 멸균이 간편한 생물배양기 및 이를 이용한 액상배양방법
CN101380467B (zh) 多联灭活疫苗病毒(抗原)液浓缩工艺
CN203462046U (zh) 用于制备胶原蛋白的酶解反应罐
Attri et al. Sterilization in bioprocesses
CN215560294U (zh) 一种微藻光发酵培养装置
CN218290430U (zh) 一种瓶装或桶装天然矿泉水无菌灌装生产系统
CN208279451U (zh) 一种多功能自动净水系统
RU2682638C2 (ru) Способ промышленного получения морской воды, подходящей для пищевого применения
CN104099288A (zh) 一种新生牛血清的生产工艺
CN208200987U (zh) 一种用于固定化乳糖酶的生物转化反应器
CN223274817U (zh) 一种枸杞原浆快速杀菌装置
TWI236344B (en) Process for the enzymatic lactose cleavage
JP2008207154A (ja) 消化液処理方法及びその装置
CN205556068U (zh) 一种多功能实验动物饮水瓶灌装设备
Djayanti et al. Quick sterilization of Spirulina powder through dry ozonization for pharmaceutical preparations
CN218515035U (zh) 一种养殖池水循环系统

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 24860569

Country of ref document: EP

Kind code of ref document: A1