WO2021021696A1 - Dispositifs et procédés de séchage et de fermentation à l'état solide de micro-organismes - Google Patents

Dispositifs et procédés de séchage et de fermentation à l'état solide de micro-organismes Download PDF

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Publication number
WO2021021696A1
WO2021021696A1 PCT/US2020/043665 US2020043665W WO2021021696A1 WO 2021021696 A1 WO2021021696 A1 WO 2021021696A1 US 2020043665 W US2020043665 W US 2020043665W WO 2021021696 A1 WO2021021696 A1 WO 2021021696A1
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WIPO (PCT)
Prior art keywords
bag
compartment
solid
substrate
drying
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Application number
PCT/US2020/043665
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English (en)
Inventor
John Joseph CASCINO
Brady BEEMER
John Dennis FOUGERE
Lauren BRADY
Michael Harrison FETHE
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Novozymes Bioag A/S
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Publication of WO2021021696A1 publication Critical patent/WO2021021696A1/fr

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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
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/24—Gas permeable parts
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B65—CONVEYING; PACKING; STORING; HANDLING THIN OR FILAMENTARY MATERIAL
    • B65D—CONTAINERS FOR STORAGE OR TRANSPORT OF ARTICLES OR MATERIALS, e.g. BAGS, BARRELS, BOTTLES, BOXES, CANS, CARTONS, CRATES, DRUMS, JARS, TANKS, HOPPERS, FORWARDING CONTAINERS; ACCESSORIES, CLOSURES, OR FITTINGS THEREFOR; PACKAGING ELEMENTS; PACKAGES
    • B65D81/00—Containers, packaging elements, or packages, for contents presenting particular transport or storage problems, or adapted to be used for non-packaging purposes after removal of contents
    • B65D81/32—Containers, packaging elements, or packages, for contents presenting particular transport or storage problems, or adapted to be used for non-packaging purposes after removal of contents for packaging two or more different materials which must be maintained separate prior to use in admixture
    • B65D81/3261—Flexible containers having several compartments
    • B65D81/3266—Flexible containers having several compartments separated by a common rupturable seal, a clip or other removable fastening device
    • 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
    • C12M21/00—Bioreactors or fermenters specially adapted for specific uses
    • C12M21/16—Solid state fermenters, e.g. for koji production
    • 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
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/14—Bags

Definitions

  • Solid-state fermentation involves cultivation of microorganisms, like fungi or bacteria, on solid or semi-solid substrates.
  • the cultivation, or fermentation is usually performed in a bioreactor. After the cultivation, material from the bioreactor may be dried, and the
  • microorganisms and accompanying substrate preserved or further separated or extracted from the substrate.
  • microorganisms are active areas of research. Development of improved devices for solid-state fermentation is of interest.
  • Solid-state fermentation of microorganisms, on moist solid or semi-solid substrates, and subsequent drying of the substrates generally requires transfer of the microorganisms/substrate from a device where fermentation occurs to a device that facilitates drying. Transfer of the microorganisms/substrate between devices can increase the risk of contaminating the microorganism cultures and can expose human operators to the microorganisms.
  • the disclosed devices have or are configurable to have compartments for fermenting microorganisms and compartments for drying microorganisms/substrates.
  • the compartments are configured so that microorganisms fermented on a substrate in one compartment can be repositioned to another compartment for drying. Repositioning of the
  • microorganisms/substrates from the fermentation compartment to the drying compartment, is performed without exposure of the microorganisms to the external environment, and without exposing human operators to the microorganisms during the repositioning.
  • the fermentation compartment of the device and the drying compartment of the device differ from one another in their permeability to water and/or water vapor. The difference in water permeability of the compartments provides controlled-rate desiccation of the substrate from both the fermentation compartments and the drying compartments of the devices.
  • Fig. 1 illustrates how an example drying bag can be shortened, and thus controlling the length, by movement of an external clip on the drying bag.
  • Fig. 2A illustrates a top view of an example solid-state fermentation and drying bag.
  • Fig. 2B illustrates a side view of the example solid-state fermentation and drying bag shown in Fig. 2A.
  • Fig. 2C illustrates another side view of the example solid-state fermentation and drying bag shown in Fig. 2A.
  • Fig. 2D illustrates another top view of the example solid-state fermentation and drying bag shown in Fig. 2A, shown here with an example clip not positioned on the bag.
  • Fig. 2E illustrates another top view of the example solid-state fermentation and drying bag shown in Fig. 2A, shown here with an example clip positioned on the bag to provide a fermentation compartment and a drying compartment.
  • Fig. 3A illustrates a top view of an example solid-state fermentation and drying bag.
  • Fig. 3B illustrates a side view of the example solid-state fermentation and drying bag shown in Fig. 3A.
  • Fig. 3C illustrates another top view of the example solid-state fermentation and drying bag shown in Fig. 3A, shown here with an example clip not positioned on the bag.
  • Fig. 3D illustrates another top view of the example solid-state fermentation and drying bag shown in Fig. 3A, shown here with an example clip positioned on the bag to provide a fermentation compartment and a drying compartment.
  • Fig. 4 illustrates a top view of an example solid-state fermentation and drying bag.
  • Fig. 5 illustrates a top view of an example solid-state fermentation and drying bag.
  • Fig. 6 illustrates a top view of an example solid-state fermentation and drying bag, where the drying bag has only one side with a filter, which has been extended in length for faster drying. It also has incorporated a small frame of plastic around all sides of the filter on the drying bag to increase strength.
  • bag means a flexible container that can be closed to the external environment.
  • clip means a device that can be applied to a bag to divide the bag into separate compartments.
  • the term “close” or“closed,” when referring to devices, may mean that the interior of the device is not open to or accessible to the external environment.“Close” or “closed” may also refer to the relationship of compartments of a device to one another and mean that a passage between the compartments is blocked or covered.
  • the term“closable” means something configured so that it can be closed.
  • the term“closure” when used as a noun generally refers to an object that, when applied to a fermentation and drying device, divides or separates an interior space of the device into separate compartments.
  • a clip may be considered a type of closure.
  • the term "compartment” means a section of a container.
  • configure means to arrange or organize in a particular way or form.
  • the term "configurable to” means able to be configured.
  • the term “container” means an object for holding or containing something.
  • a covering when used as a noun, refers to something that extends over the top of something else to block passage.
  • a covering generally refers to water-impermeable material that fits on top of water-permeable material on the surface of a device, blocking exit of water vapor from the interior of the device.
  • the term "device” means a thing adapted for a particular purpose.
  • the device is a container for solid-state fermentation and drying of microorganisms and/or substrates.
  • drying means to remove moisture or liquid from a thing.
  • the term“external environment,” in reference to a fermentation and drying device, refers to locations outside of the device.
  • the term “fermentation” means cultivation or growth of microorganisms.
  • filter material refers to materials flexible materials that are permeable to water vapor.
  • the term "irreversible" means refers to an action that produces an effect or state, where the previous state cannot be restored.
  • porous means porous to a substance.
  • the term“pliable” means able to bend without breaking; flexible.
  • open when referring to devices, may mean that the interior of the device is accessible to the external environment.“Open” or“opened” may refer to the relationship of compartments of a device to one another and mean that a passage between the compartments is not blocked or covered.
  • the term“openable” means something configured so that it can be accessed; to create a passage between.
  • the term "reposition” means to adjust the position of something; to relocate something.
  • the term “reversible” refers to an action that produces an effect or state, where the previous state can be restored.
  • the term“sealed” generally refers to a bag that was once open to the environment and now is closed to the environment. In one example, an open, unsealed end of a bag may be sealed to provide a bag that is sealed to the environment.
  • solid-state fermentation refers to cultivation of microorganisms on a solid or semi-solid substrate. This contrasts with liquid fermentation, which refers to cultivation of microorganisms in liquid medium.
  • sterile means free from living organisms (e.g.,
  • substrate refers to substances used for growth of a
  • substrates are sold or semi-solid, as opposed to liquid.
  • suitable means appropriate or sufficient for a particular purpose or situation.
  • vapor refers to the gaseous phase of a substance.
  • an all-in-one device e.g., a device used for both fermentation of microorganisms and drying of substrates
  • Some of the compartments of these devices may differ in their permeability to water/water vapor compared to other of the compartments.
  • the compartments are not open to the external environment. In general, contaminants from the environment external to the devices are not able to contaminant the microorganisms located in the interior of the devices.
  • the multiple interior compartments of the devices generally can be of two types.
  • One type of compartment is suitable for fermentation of microorganisms using a solid or semi-solid substrate (i.e. , solid-state fermentation).
  • Another type of compartment is suitable for drying of the solid or semi-solid substrate used for the fermentation.
  • the fermentation compartments and the drying compartments generally differ from one another in permeability to water and/or water vapor. Generally, the fermentation compartments are less permeable to water/water vapor than the drying compartments. That is, the drying compartments generally are more permeable to water/water vapor than the fermentation compartments.
  • This configuration generally restricts evaporation of water from a hydrated or moist microbial substrate (e.g., rice) during fermentation of microorganisms, and promotes evaporation of water from hydrated microbial substrates during drying of the fermented microorganisms.
  • a hydrated or moist microbial substrate e.g., rice
  • drying compartments may have more water-permeable vents than fermentation compartments.
  • drying compartments may be made or partially made from materials that are more permeable to water/water vapor than are the fermentation compartments. Drying compartments may be configured such that airflow into and out of (e.g., through) these compartments is greater than airflow through the fermentation compartments. In these devices, airflow through the
  • compartments may increase flow of water vapor out of the compartments.
  • moisture is more easily, more efficiently removed from drying compartments of the devices than from the fermentation compartments of the devices.
  • the devices are configured so that microorganisms fermented in a fermentation compartment can be moved or repositioned from the fermentation compartment to the drying compartment after fermentation has occurred.
  • fermentation and drying compartments may be proximate to one another.
  • fermentation and drying compartments may be adjacent to one another.
  • the fermentation and drying compartments of the device may be open to one another. In an open configuration,
  • microorganisms/substrate may be repositioned between compartments.
  • fermentation and drying compartments may be closed to one another. In a closed configuration, microorganisms/substrate may not be repositioned between compartments.
  • compartments that are open to one another may be able to be closed to one another.
  • compartments that are closed to one another may be able to be opened to one another.
  • the ability to open and close the compartments to one another may be reversible in some examples of the device.
  • the device may have a reversible closure that provides for this.
  • the ability to open and close the compartments to one another may not be reversible.
  • the devices generally are constructed in a way, and/or from materials, that can be sterilized. That is, when sterilized, the fermentation chambers of the devices are usable for fermentation of cultures of microorganisms, and the drying chambers are usable for drying of cultures of microorganisms containing moist substrate, without contamination of the cultures from the exterior of the device.
  • the devices generally may also be constructed in a way, and/or from materials, that provide for air exchange between the interior and the exterior of the device.
  • the device may contain valves that provide for, regulate, or prevent exchange of air or gas between the interior and exterior and/or between the exterior and interior of the device. Air exchange may be needed by the microorganisms during fermentation. Air exchange may facilitate drying of the microorganisms after the fermentation.
  • the devices may also be constructed in a way, and/or from materials, that provide for water and/or water vapor exchanged between the interior and exterior of the device (e.g., moisture removed from the interior of the device).
  • fermentation compartments of the devices generally are less permeable to water/water vapor than drying compartments of the devices.
  • This differential in water permeability may also be accomplished in different ways (e.g., valves, vents, materials, air flow, and the like), some examples of which are discussed here.
  • the solid-state fermentation and drying device is a bag.
  • the bag may have various shapes. In some examples, the bag may be square or rectangular. In some examples, the bag may be cylindrical. The bag may be gusseted. The bag may be of various sizes.
  • the interior of the bag may have multiple compartments or may have a compartment that can be divided into multiple compartments (e.g., fermentation compartment and drying compartment).
  • Individual compartments may be open or openable to one another.
  • Individual compartments may be closed or closable to one another.
  • the compartments may be reversibly or irreversibly openable or closable to one another.
  • bags may be used to provide the compartments and/or to open or close compartments to one another.
  • bags may be configured to have built-in closures, that may be reversible.
  • a bag may have a zipper that can be opened or closed by moving a tab.
  • a clip may be applied to a bag to create compartments in a bag that are closed to one another.
  • An example clip may push separate walls of a bag together to create compartments that are closed to one another.
  • Clips may include reversible bag sealers, one type being a GRIPSTIC® Bag Sealer.
  • Bags may have one or more openings that provide for adding substances, like substrates for solid-state fermentation, to one or more interior compartments.
  • the opening may be configured to be closed or sealed to the external environment.
  • fermentation substrate may be added to the bag through an opening, and the opening may be closed (e.g., by sealing) after addition of substrate to the bag.
  • the bags may be made from pliable materials that are not permeable to water (e.g., polypropylene, polyethylene, high-density polyethylene).
  • the bag material may be permeable or semi-permeable to air.
  • the pliable bag may be made from polypropylene material.
  • Polypropylene can be sealed using heat. Polypropylene is tolerant to high temperatures (about 256 degrees Fahrenheit), which may facilitate sterilization of the bag and its contents (e.g., substrate for fermentation).
  • the materials that comprise the bag are compatible with microorganism viability and do not inhibit microorganism growth.
  • the thickness of the material may be from 2.2 to 4.0 mil.
  • the bags may be constructed so that, when configured into interior compartments, fermentation compartments differ in their permeability to water and/or water vapor from drying compartments.
  • a relatively non-water permeable material like polypropylene, may be interspersed with a water permeable material on the surface of the bag.
  • compartment suitable for fermentation of microorganisms may be constructed to have a surface area with less water permeable material than a bag compartment suitable for drying.
  • water-permeable areas of the bag may be made using filter material.
  • filter material Various types of filter material may be used.
  • the filter material may be Tyvek®.
  • the filter material may be Unicorn Type A (e.g., 0.5 microns, laminate of non-woven polypropylene and polypropylene membrane), Type B, Type BN, Type T (0.2 microns), Type 14/14A and the like (Unicorn Imp. & Mfg. Corp., Plano, Texas, USA;
  • the filter material may have an example pore size of from 0.2 to 5 microns.
  • the filter material generally is permeable to air and water, but may not be permeable to contaminants, like mold spores and bacteria.
  • the water-permeable areas of the bag may contain one layer of filter material.
  • the water permeable areas of the bag may contain 2, 3, 4, 5 or more layers of filter material.
  • the bags and materials from which they are made are generally are sterilizable by one or more methods known in the art (steam sterilization, gamma irradiation, ethylene oxide sterilization, electron-beam radiation, STERRAD® sterilization, and the like).
  • example bags may be described as in the Drawings.
  • Fig. 1 shows how an adjustable length of the drying bag can be used to control the drying rate by manipulating surface to volume ratio in the drying bag.
  • the length of the drying bag can be shortened by movement of an external clip on the drying bag.
  • the location of the clip on the drying bag filter will determine the substrate height and can affect the substrate surface area to volume ratio. A decrease in this ratio slows the drying process and an increase speeds up the drying process. Being able to fine tune the drying rate of a specific microbe can have a positive effect on the microorganism’s survival.
  • FIG. 2A illustrates a top view of the bag 200.
  • the bag 200 pictured in these figures is made of pliable material and appears flat, with a left end 210 and a right end 220.
  • this example bag 200 is cylindrical and the two ends of the bag shown in Fig. 2A are ends 210, 220 of the flattened cylinder.
  • the sides of the cylinder of the pliable bag 200 are spread apart (in Fig. 2A they are flattened together) to better show the cylindrical shape of the bag 200.
  • the bag material is made of a substantially water-impermeable nature
  • the bag 200 contains regions of water-permeable material which, in this example, is filter material.
  • a region of water-permeable material located on the left side of the bag 200 is shown as 230.
  • the inserts of water-permeable material 240, 242 on the right side of the bag 200 are shown to be positioned on opposing sides of the cylinder that makes up the bag 200.
  • the regions, or inserts of water-permeable material 230, 240, 242 (e.g., filter material) provide for escape of water and/or water vapor to the external environment from the interior of a sealed bag.
  • a region located between the inserts of water-permeable material located on the left side of the bag 230, and the inserts of water-permeable material located on the right side of the bag 240, 242, is called a separator region 250.
  • a separator can be applied to the separator region 250 of the bag to separate the bag into different
  • Figs. 2D and Fig. 2E illustrate this.
  • Fig. 2D is a top view of the bag 200 that also shows an example separator.
  • the separator is a clip 260.
  • Fig. 2E shows the clip 260 applied to the bag 200 at the separator region 250. The clip 260 pinches the sides of the pliable bag together.
  • the bag 200 is separated into separate compartments. Neither air or water can move from one compartment to the other when the clip 260 is in place.
  • the compartment located on the left side of the bag 200 in this example is called the fermentation compartment 270.
  • the compartment located on the right side of the bag 200 in this example is called the drying compartment 280.
  • the fermentation compartment 270 contains less water-permeable material than the drying compartment 280.
  • the left end 210 of the bag 200 is sealed with the clip 260, positioned as shown in Fig. 2E, the interiors of both the fermentation compartment 270 and the drying compartment 280 are not exposed to the external environment.
  • FIG. 3A-D Another example of a solid-state fermentation and drying bag is shown in Figs. 3A-D.
  • the bag 300 shown in Figs. 3A-D is cylindrical, with a left end 310 and a right end 320.
  • the bag 300 has a region of water-permeable material 330 located on the left side.
  • the bag 300 has a different configuration of water- impermeable material and water-permeable material on the right side of the bag, as compared to the bag 200 illustrated in Figs. 2A-E.
  • the regions of water-permeable material 240, 242 on the right side of the bag are interspersed with water- impermeable plastic bag material (i.e. , the regions 240 and 242 may be said to be“inserts”).
  • the bag 300 illustrated in Figs. 3A-D has water-permeable material (e.g., filter material) that, in this example, essentially encompasses the entire right side of the bag.
  • the part of the bag to the right of the separator region 350 of the bag 300 is filter material.
  • substantially the entirety of the drying compartment 380 that is created by application of the clip 360 is permeable to water and/or water vapor.
  • the bag 400 has a left end 410 and a right end 420. Inserts of water-permeable material 530 are shown on the left side of the bag 400. The right side of the bag 400 is entirely made from water-permeable material.
  • the bag 400 has a clip 460 attached so that there is a fermentation compartment 470 and a drying compartment 480. Substantially the entirety of the surface of the drying compartment 480 is water-permeable, filter material.
  • the bag 500 has a left end 510 and a right end 520. Inserts of water-permeable material 530 are shown on the left side of the bag 500. The right side of the bag 500 is entirely made from water-permeable material.
  • the bag 500 has a clip 560 attached so that there is a fermentation compartment 570 and a drying compartment 580. Substantially the entirety of the surface of the drying compartment 580 is water-permeable, filter material.
  • the drying bag has only one side with a filter, which has been extended in length for faster drying. It also has incorporated a small frame of plastic around all sides of the filter on the drying bag to increase the strength of the seams.
  • the methods disclosed here relate to using the disclosed devices.
  • solid/semi-solid substrate is placed into the device.
  • the device is then sterilized under conditions such that the substrate contained within the device is also sterilized.
  • the system is then ready for cultivation of pure cultures of microorganisms.
  • the substrate within the device can then be inoculated with a culture of microorganisms to be propagated, and cultivation/fermentation of the microorganisms can be performed.
  • fermentation of microorganisms generally occurs in a compartment of the device that is suitable for fermentation (i.e. , the compartment is constructed so that evaporation of water from the substrate is restricted).
  • the substrate, including the fermented microorganisms generally are repositioned to a compartment of the device that is suitable for drying the substrate (i.e., the compartment is constructed so that evaporation of water from the substrate is favored).
  • the fermentation and drying device may be a bag.
  • a bag used in the methods is shown in Figs. 2A-E.
  • substrate for solid-state fermentation of microorganisms e.g., rice, bran
  • the open end of the bag 200 may then be sealed, or otherwise closed, so the interior of the bag, and the bag’s contents, are not exposed to the exterior environment.
  • the substrate may be positioned in the bag in various ways at this point. For example, referencing Fig.
  • the substrate may be placed in the part of the bag 200 that, after application of a clip 260, is referred to as the fermentation compartment 270.
  • the substrate may be placed in the part of the bag 200 that, after application of a clip 260, is referred to as the drying compartment 280.
  • the substrate may be placed in the bag 200 such that substrate is located in both the fermentation compartment 270 and the drying compartment 280.
  • the clip 260, or other component used to divide the bag 200 into separate compartments may be in position on the bag 200 (as in Fig. 2E) or may not be positioned on the bag 200 (as in Fig. 2D).
  • the sealed bag is then sterilized in a way that the substrate contained within the bag is also sterilized.
  • a variety of methods known in the art may be used for sterilization, including for example, steam sterilization, gamma irradiation, ethylene oxide sterilization, electron-beam radiation, STERRAD® sterilization, and the like.
  • the substrate may be repositioned there.
  • the bag shown in Figs. 2A-E it may be possible to raise one end of the bag to cause substrate in the bag to move to the other end of the bag due to gravity.
  • the fermentation may be performed using the bag shown in Figs. 2A-E.
  • Fig. 2E may then be closed to the drying compartment, by applying the clip 260, as shown in Fig. 2E, for example.
  • the sterile substrate in the bag is ready to be inoculated with a culture or cultures of microorganisms to be cultivated. Inoculation may be performed in a variety of ways.
  • the bag may have been constructed to contain one or more ports or other openings in the bag, that provide for inoculation without risk of contamination.
  • a syringe containing an inoculum may be prepared. A hypodermic needle attached to the end of the syringe may be used to puncture the surface of the bag and inject the inoculum in the syringe.
  • the bag is placed in an environment (e.g., temperature, humidity, and the like) that is conducive for growth of the microorganisms.
  • an environment e.g., temperature, humidity, and the like
  • the substrate is repositioned from the fermentation compartment to the drying
  • the clip 260 is removed from its position on the bag 200, to open the fermentation compartment to the drying compartment.
  • the substrate, located in the fermentation compartment 230 is then repositioned to the drying compartment 280, without opening the bag to the external environment.
  • repositioning can be done by elevating the left end 210 of the bag 200 above the right end 220 of the bag 200, so that the substrate and microorganisms slide from the fermentation compartment 270 to the drying compartment 280, because of the force of gravity.
  • the bag can be placed in an environment that is conducive for drying of the substrate.
  • the clip 260 may remain off the bag 200, or the clip 260 may be repositioned on the bag 200, as shown in Fig.
  • the bag 200 may be opened to the external environment and the
  • microorganisms removed.
  • the microorganisms may be extracted from the substrate.
  • a device for solid-state fermentation of microorganisms, and for drying of the solid/semi-solid substrates on which the microorganisms propagate may not have multiple interior compartments.
  • These example devices may have interior compartments that are generally suitable for both fermentation of microorganisms using a solid or semi-solid substrate and for drying of the solid or semi-solid substrate used for the fermentation.
  • water-permeability of the interior compartments of these devices may be adjustable or modifiable.
  • a fermentation and drying device is a bag that may be made of water-impermeable material (e.g., polypropylene) that is interspersed on the surface of the bag with water-permeable material (e.g., filters).
  • water-impermeable material e.g., polypropylene
  • at least some of the water-permeable material may be covered by water-impermeable or less water-permeable material (e.g., polypropylene may cover the surface of the filter material).
  • water/water vapor attempting to leave the interior of the bag through the water-permeable material may be blocked or restricted by the water-impermeable material covering the water-permeable material.
  • fermentation of microorganisms would generally be conducted with at least some of the water-permeable material covered to prevent/slow desiccation of moist solid/semi-solid substrate in the bag during the fermentation.
  • some or all the water-impermeable material covering the water-permeable material is removed, to increase the ability of water/water vapor to leave the interior of the bag during drying (i.e. , water vapor leaving the bag through the water-permeable material would not be impeded).
  • the bag is placed under conditions that promote drying of the substrate in the bag.
  • water-permeable filter material e.g., filters
  • bag material e.g., polypropylene
  • plastic zippers e.g., Easy Open Tabs found in ZIPLOC® bags
  • VELCRO® e.g., VELCRO®
  • the polypropylene material may be“zipped off’ or“ripped off” to provide increased water/water vapor exchange through the filter material.
  • the material used to cover the water-permeable filter material may be paper or plastic with adhesive on one surface.
  • the adhesive paper/plastic may be adhered over the filter material during fermentation, to prevent or reduce evaporation of moisture from the bag.
  • the paper/plastic may then be removed (e.g., pulled off) after fermentation, to facilitate drying of the substrate.
  • a fermentation bag system may include two bags, one that fits inside of the other (bag-in-a bag design).
  • the smaller, interior bag is suitable for fermentation of microorganisms on a solid/semi-solid substrate.
  • the smaller bag generally is more permeable to water/water vapor than the larger bag, into which the smaller bag fits (the exterior bag).
  • fermentation is designed to occur within the smaller bag, while the smaller bag is located inside of a sealed, larger bag.
  • the larger bag is designed to be less permeable to water/water vapor than the smaller bag (i.e. , water evaporation from the system is limited by water permeability of the outer bag), evaporation of water from the fermentation can be limited during fermentation.
  • both bags may be made of water- impermeable material (e.g., polypropylene) that is interspersed with water-permeable material (e.g., filters).
  • the inner bag may contain relatively more water-permeable material on its surface than the exterior bag.
  • permeability to water during fermentation is controlled by water-permeability of the outer bag.
  • Permeability to water during drying is controlled by water- permeability of the inner bag, which has by that time been removed from the outer bag.
  • a clip can be used to isolate nutrients/substrate or other fermentation fed-batch components typically used in liquid fermentation (i.e., sucrose, yeast extract, minerals, salts, or buffers) from the growth side of the fermentation bag.
  • this clip may be removed to transfer per determined amounts of components as a bolus to the actively growing fermentation.
  • This addition will improve the growth rate of the organisms being cultivated. It may be used as an osmotic stress to improve desiccation survival of the organisms under a solid-state fermentation setting.
  • it may enhance the length of fermentation time with the addition of nutrients and buffer allowing inert substrates to be replenished with nutrients consumed by the organism being grown.
  • Another implementation of this technology is to alter the pH of the growth substrate to either initiate sporulation or prolong the fermentation to improve fermentation yields. This technology is commonly used in liquid fermentation, but not in a solid-state enclosed fed-batch system.
  • adding a drying agent such as calcium carbonate, calcium oxide, calcium sulfate, zeolite, water absorbing crystals made of sodium polyacrylate, silica gel, activated carbon, and the like, to the drying side of the two-chambered bag would speed up drying.
  • the moisture content would immediately be lowered by mixing wet substrate with a lower moisture content drying agent.
  • the drying agent would accelerate further drying of the substrate by accelerating the absorption of moisture from the wet fermented substrate.
  • the drying agent could be sterilized in the drying chamber at the same time the nutritive substrate is sterilized on the fermentation side of the bag. This would ensure the drying agent is sterile when the wet ferment is transferred to the drying side of the bag. Drying agents would be in an initial moisture range under 10% and more preferably 1-5%.

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Abstract

L'invention concerne des dispositifs pour la fermentation à l'état solide de microorganismes et le séchage de microorganismes produits par fermentation à l'état solide. L'invention concerne également des procédés d'utilisation des dispositifs.
PCT/US2020/043665 2019-07-30 2020-07-27 Dispositifs et procédés de séchage et de fermentation à l'état solide de micro-organismes WO2021021696A1 (fr)

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11066632B1 (en) 2021-03-31 2021-07-20 Wei K. Hsu Cultivation bag assembly for cultivating microbes
US11178826B1 (en) 2021-03-31 2021-11-23 Wei K. Hsu Cultivation bag assembly for cultivating microbes
USD937094S1 (en) 2021-04-22 2021-11-30 Wei K. Hsu Dual cultivation bag
CN113911538A (zh) * 2021-10-12 2022-01-11 安徽宸瑞医药包装有限公司 一种多层无菌收纳袋
USD961801S1 (en) 2021-11-11 2022-08-23 Wei K. Hsu Incubation bag

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5836445A (en) * 1996-07-23 1998-11-17 Fmc Corporation Pouch
WO2004111181A1 (fr) * 2003-06-13 2004-12-23 Elep S.A.S. Di Cabiddu Rachele & C. Installation pour fermentation a l'etat solide
EP2505634A1 (fr) * 2011-03-31 2012-10-03 Technelep srl Installation pour la fermentation en milieu solide
WO2015180908A1 (fr) * 2014-05-28 2015-12-03 Ge Healthcare Bio-Sciences Ab Ensemble sac pour la culture de cellules
US20160023834A1 (en) * 2010-06-17 2016-01-28 David DiLiberto Multi-compartment container with frangible seal and vapor permeable region
EP3677666A1 (fr) * 2019-01-02 2020-07-08 Wei K. Hsu Sac de culture pour cultiver des microbes

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5836445A (en) * 1996-07-23 1998-11-17 Fmc Corporation Pouch
WO2004111181A1 (fr) * 2003-06-13 2004-12-23 Elep S.A.S. Di Cabiddu Rachele & C. Installation pour fermentation a l'etat solide
US20160023834A1 (en) * 2010-06-17 2016-01-28 David DiLiberto Multi-compartment container with frangible seal and vapor permeable region
EP2505634A1 (fr) * 2011-03-31 2012-10-03 Technelep srl Installation pour la fermentation en milieu solide
WO2015180908A1 (fr) * 2014-05-28 2015-12-03 Ge Healthcare Bio-Sciences Ab Ensemble sac pour la culture de cellules
EP3677666A1 (fr) * 2019-01-02 2020-07-08 Wei K. Hsu Sac de culture pour cultiver des microbes

Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11066632B1 (en) 2021-03-31 2021-07-20 Wei K. Hsu Cultivation bag assembly for cultivating microbes
US11178826B1 (en) 2021-03-31 2021-11-23 Wei K. Hsu Cultivation bag assembly for cultivating microbes
USD937094S1 (en) 2021-04-22 2021-11-30 Wei K. Hsu Dual cultivation bag
USD947680S1 (en) 2021-04-22 2022-04-05 Wei K. Hsu Dual cultivation bag
CN113911538A (zh) * 2021-10-12 2022-01-11 安徽宸瑞医药包装有限公司 一种多层无菌收纳袋
CN113911538B (zh) * 2021-10-12 2023-04-28 安徽宸瑞生物科技有限公司 一种多层无菌收纳袋
USD961801S1 (en) 2021-11-11 2022-08-23 Wei K. Hsu Incubation bag

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