RU2363729C1 - Apparatus for suspension cultivation of tissue or microorganism cells - Google Patents

Abstract

FIELD: microbiology instruments.

SUBSTANCE: invention relates to suspension cultivation of tissue or microorganism cells and can be used in food industry, biotechnologies and medicine. Proposed apparatus comprises cylindrical vessel with cover and device to aerate and mix suspension of mentioned biological substances. The said apparatus allows producing, on suspension surface, aerating gas swirled flow flowing along the vessel edges, potential vortex rate field being located at the vessel edges and axial countercurrent in near-axis zone. Note that the pressure difference between the vortex edges and centre varies from 10 to 2000 Pa. Axisymmetric body of revolution is arranged inside the said vessel, on its bottom and aligned with it, the body height being equal to or exceeding the level of the apparatus maximum filling. Side surface of aforesaid body of revolution smoothly mates the vessel bottom surface. Aforesaid body of revolution represents a cone or a cylinder with aerodynamic nose fairing arranged on its upper end face.

EFFECT: higher efficiency of cultivation due to more regular distribution of cless culture in fluid substrate.

2 cl, 4 dwg

Description

The invention relates to devices for suspension cultivation of tissue cells or microorganisms and can be used in the food industry, biotechnology and medicine.

Known bioreactor for the cultivation of cell cultures on microcarriers under microgravity conditions (US patent No. 5002890, IPC C12M 3/06, publ. 26.03.1991). The bioreactor contains a vertical axisymmetric chamber in which a filtering device is arranged coaxially with rotation, around which flexible membranes are arranged to rotate. Liquid nutrient medium enters the chamber with cells immobilized on microcarriers through a filtering device from a closed system for preparing the nutrient medium and its aeration. The aeration of the nutrient medium with gaseous components is perfused through a semipermeable membrane. The release of the used medium is carried out in the same closed system. In a closed system, the input and output parameters are read by sensors, through which nutrients are added and pH is regulated, oxygen is also supplied, carbon dioxide is removed and gas bubbles are eliminated. The specified system is under the control and control of the microprocessor. This bioreactor design is designed to work in zero gravity.

However, such a device is difficult both constructively and during operation and has low mass transfer characteristics due to the perfusion method of aeration of the cell suspension.

The closest analogue (prototype) is an apparatus for suspension cultivation of tissue cells or microorganisms (PCT Application No. 92/05245, IPC C12M 1/04, publ. 02.04.92), containing a cylindrical container with a lid and nozzles for supplying and discharging gas and a device for aeration and mixing of the medium. The aeration and mixing device comprises a horizontal impeller mounted on a vertical drive shaft, located in the upper part of the container directly under the cover, and an annular plate located underneath with a central gas outlet, peripherally attached to the container wall to form an annular cavity around the wheel for supply and removal of gas. Slit openings for gas passage are made in the annular partition, arranged uniformly around the circumference at an angle to the horizontal plane. A gas supply pipe is installed in the lid coaxially with the impeller, and a gas pipe is connected to the specified annular cavity and is located on the edge of the cover.

However, the prototype has the following disadvantages. Due to the law of conservation of angular momentum, the vortex flow in the gas and liquid consists of a vortex ring located in the interval from the axis of the vessel to its side walls, and a vortex “cord” located on the axis of the vessel. The complex vortex nature of the flow of the substrate leads to the fact that, according to the laws of hydrodynamics, some of the cells begin to concentrate in the central vortex cord of the substrate. As a result of this uneven distribution of cells or microorganisms by the volume of the liquid substrate, the delivery process of oxygen to the cells weakens, which leads to a decrease in the efficiency of the process of cultivation of aerobic microorganisms.

The technical result of the invention is to increase the efficiency of the cultivation of cell cultures or microorganisms by providing a more uniform distribution of cell cultures or microorganisms in the volume of a liquid substrate.

The specified technical result is achieved by the fact that in the apparatus for suspension cultivation of tissue cells or microorganisms containing a cylindrical container with a lid and a device for aeration and mixing of the suspension of these biological objects, made with the possibility of creating on the surface of the suspension a swirling stream of aerating gas with a potential vortex velocity field of the periphery of the tank and the axial countercurrent in the axial zone and the pressure drop between the periphery and the center of the vortex within 10-2000 Pa, co According to the invention, an axially symmetric body of revolution is installed in the vessel coaxially with the latter, which is attached with an end to the bottom of the vessel, the height of which is equal to or greater than the level of maximum filling of the apparatus’s capacity, and the lateral surface of this rotation body is smoothly mated with the surface of the vessel’s bottom.

The body of rotation is made in the form of a cone or in the form of a cylinder with an aerodynamic fairing located in its upper end.

In the proposed design of the apparatus for vortex suspension cultivation of tissue cells or microorganisms in comparison with the prototype apparatus, the occurrence of a vortex cord in a liquid substrate is excluded and thereby the cultivation conditions of aerobic microorganisms are improved due to their more uniform distribution in suspension.

A brief description of the drawings. In the following, the invention is illustrated by a detailed description of examples of its implementation with reference to the accompanying drawings, in which: Fig. 1 shows a schematic general view of an apparatus for suspension cultivation of tissue cells or microorganisms with a conical body of revolution in a container; figure 2 is the same with a cylindrical body of revolution; figure 3 shows a video frame with visualization of the vortex flow in a liquid with particles in the design of a bioreactor with a flat bottom (prototype); figure 4 shows a video frame with visualization of the vortex flow in a fluid with particles in the design of the inventive bioreactor with a conical insert and a rounded bottom of the vessel with a liquid substrate.

Embodiments of the invention

The apparatus for suspension cultivation of tissue cells or microorganisms contains a cylindrical container I (Fig. 1) for suspension of cells with a cover 2 and a pipe 3 for supplying aerating gas and venting a gaseous medium and a device 4 for mixing a suspension of cells, said device 4 and pipe 3 being placed in the upper part of capacity 1 above the level aa of the cell suspension in this capacity. The device 4 for mixing and aeration of the cell suspension is configured to create in the upper part of the container I above the surface of the cell suspension a swirling stream of aerating gas with a potential vortex velocity field at the periphery of the container I and an axial countercurrent in the axial zone and a pressure drop between the periphery and the center of the vortex within 10-2000 Pa.

In accordance with figure 1, the device 4 for mixing the cell suspension is made in the form of a blade wheel 5 mounted for rotation on the shaft 6 by means of an electric motor 7. In the lower part of the tank I (in figure 1) there is a pipe 8 for introducing the culture medium and seed . The same pipe 8 also serves to drain the cell suspension after the end of the cultivation process.

An axially symmetric body of revolution 9 is mounted coaxially with the latter in the vessel 1, which is attached with an end to the vessel bottom, the height of which is equal to or greater than the maximum filling level of the apparatus vessel, and the side surface of the indicated rotation body smoothly mates with the surface of the vessel bottom 1.

The body of rotation is made in the form of a cone, or in the form of a cylinder with an aerodynamic fairing 10 installed in its end. Above level aa of the filling of the cell suspension, a gas cavity 11 is formed.

The work of the proposed apparatus is as follows.

In sterile conditions, the cylindrical container I is filled with a nutrient medium so that a cavity 11 for aerating gas remains above the surface of the cell suspension in the upper part of the container 1. Then turn on the temperature regulator for the desired temperature regime of cultivation and enter the seed dose of cells. Further include a device 4 for mixing and aeration of the cell suspension, which creates a quasistationary rotational motion in the cell suspension generated by a vortex flow of aerating gas. To do this, include an electric motor 7, which rotates the impeller 5. When the impeller 5 rotates, a vacuum is created in its axial region and increased pressure on the periphery. Under the action of a pressure drop (within 10-2000 Pa) between the periphery of the vessel and the axial zone of the cavity 11 created by the impeller 5, the gas circulates along spiral paths with high speed over the surface of the suspension with the velocity field of the potential vortex at the periphery of the vessel and an axial countercurrent in the axial zone. Depending on the requirements of the technology, the required speed of the impeller is set.

Thus, a swirling flow of aerating gas is formed over the surface of the cell suspension with a potential vortex velocity field at the periphery of the container and an axial countercurrent in the axial zone, which generates a similar turbulent quasistationary rotational motion in the liquid with intensive mixing along the axis of the container.

As a result of the intensive swirling of the aerating gas flow over the surface of the cell suspension in the vortex, there is a pressure difference between its periphery (increased) and the center (lowered). The pressure difference in the gas through the free surface of the cell suspension generates axial motion in the latter: descending in the peripheral zone of the tank 1 and ascending in its axial zone. At the same time, due to the friction of the gas on the free surface of the liquid medium, a rotational motion of the suspension occurs. During cell cultivation, the aeration gas interacts with the cell suspension through its free surface, saturating it with oxygen and not mixing with it. As a result, the formation of gas bubbles in the cell suspension is eliminated, which reduces cell injury and foam formation. Since the specified vortex mixing of the suspension is a quasistationary process, it provides the intensification of mass transfer characteristics without stagnant zones in the volume of the tank. The axisymmetric body of revolution 9, made in the form of a cone (Fig. 1) or cylinder (Fig. 2), attached to the bottom of the container 1, excludes the appearance of a central vortex cord in the cell suspension, which contributes to the concentration of cultured cells or microorganisms in the axial zone of the container 1 In this case, the flow of the liquid substrate in the form of a vortex ring is preserved.

Figure 3 shows an example (video frame) of the visualization of the process of mixing and aeration on a laboratory model of a vortex bioreactor prototype with a flat bottom. Small particles were used as an impurity for visualizing the flow, and water was used as a liquid. Figure 3 shows a video frame, clearly showing that the particles visualizing the flow are concentrated in the region of the vortex cord in the substrate.

Figure 4 shows an example (video frame) of the visualization of the flow on a laboratory model of the inventive vortex apparatus with a conical insert (axisymmetric body 9). Figure 4 shows that the particles visualizing the flow are evenly distributed over the volume of the liquid substrate in the tank 1. The claimed technical solution eliminates the disadvantage of the prototype, which consists in the negative impact of the vortex cord in the substrate on the efficiency of cultivation of aerobic microorganisms. In this case, the conical insert (body 9) and the rounded bottom of the container 1 exclude the appearance of a central vortex cord in the liquid substrate. In this case, the flow of the substrate in the form of a vortex ring is preserved.

Thus, the claimed design features of the proposed device options allow, in comparison with the prototype device, to eliminate the occurrence of a vortex cord in a liquid substrate and thereby improve the conditions for aeration and cultivation of aerobic microorganisms due to their more uniform distribution in suspension.

Claims (2)

1. Apparatus for suspension cultivation of tissue cells or microorganisms, containing a cylindrical container with a lid and a device for aeration and mixing a suspension of these biological objects, made with the possibility of creating on the surface of the suspension a swirling stream of aerating gas with a velocity field of the potential vortex at the periphery of the container and an axial countercurrent in the near-axis zone and the pressure drop between the periphery and the center of the vortex within 10-2000 Pa, characterized in that in the vessel coaxially installed the latter but the axisymmetric body of rotation, an end face attached to the bottom of the container, the height of which is equal to or greater than the maximum fill level of the container unit, and the side surface of said body of revolution smoothly joined to the bottom surface of the vessel. 2. The apparatus according to claim 1, characterized in that the body of rotation is made in the form of a cone, or in the form of a cylinder with an aerodynamic fairing located in its upper end.

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