SI24338A - Disposable bioreactor system with stirrer and mechanical antifoaming agent - Google Patents

Abstract

The invention relates to a disposable bioreactor in which we can cultivate and produce cells and viruses and their components. The disposable bioreactor is equipped with a liquid ventilator and a pneumatic propulsion turbine on the inside of the top of the vessel. The rotating turbine drives one or more mixers on the same axis and mechanically breaks the resulting foam. The turbine on the inside of the top of the container drives the flow of gas. The air entering the disposable bioreactor is guided through the wind turbine through the air outlet at the top of the vessel, which allows the turbine blades to rotate at a high speed, thereby mechanically breaking the foam formed and driving the axis of the mixer. In the present invention, the disposable bioreactor introduced with a stirrer and a mechanical anti-inflammatory agent solves the problem of an effective material transfer of gas into the liquid and the problem of foaming resulting from the intensive ventilation of the bioprocess slurry in disposable bioreactors.

Description

The invention relates to a disposable bioreactor system with a turbine attached to the inside of the top of the tank, which is driven by the blown gas and which serves to simultaneously break the foam and rotate the axis.

BACKGROUND OF THE INVENTION

Bioreactors are systems designed to cultivate organisms and viruses or their components. A disposable bioreactor is technically a reaction vessel and / or bag made of artificial materials (polyethylene, polystyrene, polytetraflorethylene, polypropylene, or ethylene vinyl acetate) that can be destroyed after use (eg incinerated) and approved by supervisory agencies such as Food and Drug Administration, FDA and European Medical Agency, EMA (Eibl R. and Eibl D., 2006). Modern bioreactors designed for the cultivation of microorganisms and mammalian cells and their components allow monitoring of macroprocessor parameters, require low energy input, substrate losses during the bioprocess are small and allow the transfer of the bioprocess to a larger scale. Unlike conventional bioreactors that need to be cleaned and sterilized before and after use, disposable bioreactors do not require purification and sterilization before and after use, which offers a unique advantage in terms of the required preparation time, chemicals and use of the bioreactor.

In the following, the term "bioprocess" will refer to all types, both aerobic and anaerobic, of cultivation in a nutrient medium. Cultivation may include the cultivation of prokaryotic cells (eg, archaea and bacteria) or eukaryotic cells (yeast, fungi, mold, plant or animal cells), genetically modified (micro) organisms or virus-infected cells. The term also includes carriers on and / or in which (micro) organisms are attached.

Hereinafter, the term "container" will refer to a confined space, compact or flexible in shape, to which a nutrient medium may be placed. Preferably, the container has a bottom, walls and top and is made of one or more components. In case the container consists of several parts, the bottom of the wall has a bottom and an upper lid, which allows the container to be closed.

- 2In the following, the term "turbine" will refer to a rotary engine that receives energy from the gas flow and converts it into useful work. The turbine consists of an axle with a housing to which the blades and ducts between the blades are attached. The gas entering the duct between the blades drives the rotation of the blades on the housing around the axle, enabling useful work which, for the purpose of the present invention, e.g. rotation of an axle to which one or more mixers are attached and mechanical breaking of the foam that comes into contact with the turbine blades.

Hereinafter, the term "pneumatic actuator" will refer to the propulsion of a directional gas flow arising from pressure changes in the gas in the vessel. For the purpose of the present invention, a pneumatic drive of gas directed to the turbine blades permits rotation of the turbine and the axis to which one or more agitators are attached.

Hereinafter, the term "mixing" will refer to an operation to achieve a uniform composition, temperature or any other property in the medium, or to accelerate the material or thermal transfer in the medium. For the purposes of this invention, the term mixing refers to the mechanical, pneumatic or fluid mixing of liquids.

In the following, the term "mixer" will refer to blades which, by rotation, allow the fluid to be stirred to achieve mixing of the material in the container. Different types of mixers are used depending on the viscosity of the material in the container and the available mixing time. Common mixers are Rushton or disc turbines, turbines with straight, sickle or curved blades, miscellaneous propeller or blade mixers.

Hereinafter, the term "foaming" will refer to the process by which foam bioprocessing is formed on the surface.

There are inventions and solutions for disposable bioreactors that can be divided according to the principle of mixing bioprocess broth. Depending on the principle of mixing, the described solutions of disposable bioreactors can be divided into 4 groups: wave-based bioreactors, shaking bioreactors, agitator bioreactors, and intake air jet bioreactors.

Single-wave bioreactors with waveguide mixing enable efficient cultivation of cell cultures by providing mixing at low shear. In comparison with

- 3Conventional bioreactors carry out normal cell cultivation in a fluctuating bag. Such types of disposable bioreactors are e.g. described by Buevink M. et al., Method and apparatus for cultivating wholly utilizing wave motion (EP 1739165), Singh V., Tree dimensional disposable bioreactor (US 2010/0203624), Niazi SK, Single-use Stationary Bioreactors and Mixing Vessels (US 2011/0217767) and Baskar D. et al., Methods for culturing eukaryotic celites (US 20120329151). Single-wave bioreactors, as described, allow for the efficient cultivation of cell cultures, and, compared to conventional reactors, for growing fast-growing cultures, there is a limitation in achieving high cell density during cultivation, mainly due to insufficient substrate ventilation (Eibl et al. ., 2010).

Disposable bioreactors in which agitation is based on shaking are intended for batch cultivation of microorganisms and cell cultures. This type of disposable bioreactor in which shaking is based on shaking was invented and described by Rubenberger K, Bioreactor with vortex mixing chamber (US 6168949). In shaker-based bioreactors, the bioprocessing broth is mixed with the horizontal movement of the bioreactor vessel, typically around the central axis. Bioreactors of this type do not perform well and show limitations in achieving high cell density during the cultivation of organisms in a viscous medium and in the cultivation of fast-growing cultures, mainly due to insufficient substrate ventilation (Eibl et al., 2010)

Disposable bioreactors with inlet gas jet mixing are newer and less represented on the market. The systems are optimized for the cultivation of microbial and cell cultures. A disposable bioreactor in which mixing takes place with an inlet gas hammer was invented and described by G. J. Zeikus, Pneumatic Bioreactor (EP 1951415 BI). Today, the main constraints on the widespread use of SUB by mixing with an inlet gas jet are due to lesser known cultivation parameters.

Unlike the bioreactors mentioned above, a disposable bioreactor is usually equipped with one or more mixers on the same axis, allowing the mixing to achieve good dispersion of air bubbles at the aerator and to create homogeneous conditions in the bioreactor vessel. The agitator bioreactor is therefore equipped with one or more identical or different agitators on the same axis. Common mixers are Rushton or disc turbines, flat, sickle or curved blades, propeller or blade mixers. Turbine mixers

generate the radial flow of the bioprocessing broth, and the propeller the axial flow. Combinations of turbine and propeller mixers on the same axis are repeatedly made, which enables good dispersion of air bubbles at the ventilator and homogeneous conditions in the bioreactor vessel. The mixing bioreactor is certainly the most appropriate system when a biological active substance manufacturer wants to replace a conventional conventional mixing bioreactor with a system based on the use of a pre-prepared bioreactor plastic container or disposable bag. Therefore, the single-use mixer bioreactors are the largest on the market. Similarity to conventional mixing bioreactors is also a significant advantage of these types of disposable bioreactor systems. Although the geometric ratios are in some cases practically identical to conventional mixing bioreactors, the volumetric oxygen leakage parameters and mixing speeds are much lower in the case of disposable systems than conventional stainless steel or toughened glass bioreactors. The key reasons for the lower volumetric oxygen permeability and mixing speeds lie in the specific purpose of disposable bioreactors as well as the properties of the container and its components. In a disposable mixing bioreactor where the actuator is mounted above or below the bioreactor vessel and the shaft passes through the seals into the vessel, the highest risk of contamination for the planned bioprocess is posed. The sterile operation of bioreactors is one of the basic requirements of any bioprocess. According to the most common causes of problems in providing sterile operation, the latter have been successfully circumvented by the use of magnetic stirrers. The use of a magnet in a mixer in a disposable bioreactor was invented and described by Carli KB, Disposable Whole Culture Vessel (US 6670171), and recently by Niazi SK, Pneumatically Agitated and Aerated Single Use Bioreactor (US 20120252108) described by a pneumatic bioreactor A propulsion that allows simultaneous mixing and ventilation of the bioprocessing slurry so that the intake air through the wind turbine enters the tank through the ventilator and simultaneously drives the agitator axis. Said disposable bioreactor, as Niazi states, allows more efficient ventilation and mixing of the bioprocessing broth, but does not solve the foaming problem that occurs in the container as a result of intensive ventilation and mixing.

Although, as mentioned above, single-use bioreactors with mechanical agitator blending are the largest on the market, none of the known solutions simultaneously prevents the foaming problem between efficient ventilation and blending of bioprocess broth.

The present invention invents and describes a disposable bioreactor that solves the problems and disadvantages of the prior art, especially by preventing the foaming problem while intensively stirring and ventilating.

The object of the invention is achieved by a bioreactor system with a turbine attached to the inside of the top of the tank, which is driven by the blown gas and which serves to simultaneously break the foam and rotate the axis that is part of the turbine, and after use the bioreactor system can be completely destroyed, e.g. with incineration.

Most preferred embodiments of the invention are disclosed in the claims.

REFERENCES

Eibl R., Kaiser S., Lombriser R., Eibl D. Disposable bioreactors: current state-of-the-art and recommended applications in biotechnology. Appl. Microbiol. Biotechnol. 2010 Mar; 86 (l): 41-9.

Eibl R., Eibl D. Single-use technology in biopharmaceutical production. Wiley, 2011

EP 1739165

US 2010/0203624

US 2011/0217767

US 20120329151

US 6168949

EP 1951415 BI

US 6670171

US 20120252108

SUMMARY

The invention relates to a disposable bioreactor in which cells and viruses and their components can be grown and produced. The disposable bioreactor is equipped with a liquid ventilator and a pneumatically powered turbine on the inside of the top of the tank. The turbine by rotation drives one or more mixers on the same axis and mechanically breaks the foam formed. The turbine on the inside of the top of the tank is driven by a gas stream. Turbine blade rotation at the same time

- 6mechanically breaks the foam formed and rotates the axis to which one or more mixers are attached. The air entering the disposable bioreactor is guided through a wind turbine through the air outlet at the top of the tank, allowing the turbine blades to rotate at high speed, mechanically shattering the resulting foam and propelling the agitator axis. The invention presents a disposable bioreactor with a mixer and a mechanical foam solver that solves the problem of efficient material transfer of gas to liquid and the foaming problem that results from the intensive ventilation of bioprocess broth in disposable bioreactors.

The invention discloses a disposable bioreactor with a mixer and mechanical foam, surprisingly, it allows the introduction of air into the liquid phase and mixing to such an extent that good dispersion of air bubbles and mixing in the bioprocessing substrate is achieved, thus creating a large contact surface between the gas and the liquid. but also a high volumetric oxygen transfer coefficient, preventing excessive foaming. The invented bioreactor, as presented in the description with accompanying drawings and claims, enables the cultivation and production of fast-growing cells and viruses.

DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a cross-section of a disposable bioreactor system with a mixer and a mechanical antifoam.

FIG. 2 shows a side view of a disposable bioreactor system with a mixer and a mechanical antifoam.

FIGURE 3 shows a top view, FIGURE 4 cross-section from above in position A and FIG. 5 cross-section from above in position B, as indicated in FIG. 1 of a disposable bioreactor system with a mixer and mechanical antifoam.

6 is a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam.

FIG. 7 shows a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam, with the turbine being driven by a gas blown into the bioprocess broth.

• ·

- 7 FIG. 8 shows a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam, with the turbine being driven by gas blown into the gas phase of the bioreactor system.

FIG. 9 shows a top view of a disposable bioreactor system with a mixer and a mechanical antifoam, with the turbine being driven by gas blown at several places into the bioreactor system. FIG.

FIGURE 10 shows a side view of a disposable bioreactor system with a mixer and mechanical antifoam attached to the lid of the bioreactor vessel.

FIG. 11 shows a side view of a disposable bioreactor system with a mixer and a mechanical antifoam with a conical shaped top of the container.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a disposable bioreactor with agitator and mechanical antifoam in which cells and viruses and their components can be grown and produced.

Figure 1 shows a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam consisting of a container 1, a turbine 2 that is secured via the top slider 12 to the inside of the top of the container 1, an axis of the mixer 3 that is attached to a turbine 2, a mixer 4 attached to the axis 3, and a liquid ventilator 5. A rotary pneumatically actuated turbine 2 drives one or more mixers 4 on the same axis 3 while mechanically breaking the foam formed. The pneumatically-driven turbine 2 is driven by a gas stream blown through tubes 7, 8. The tubes have air flow regulators 9, 10 in place, allowing for regulated air intake, thereby providing for regulation of the bioprocessing slurry 6 through the aerator 5 and / or rotational speed turbine 2. Rotation of the blades 2a of the turbine 2 simultaneously mechanically breaks the foam formed from the bioprocess broth 6 into the gas phase space 20 and rotates the axis 3 to which one or more mixers are attached 4. The air entering the tube 7 enters the the gas phase and the air entering the tube 8 into the liquid phase in the disposable bioreactor and exiting via the wind turbine 2 through the air outlet 11 at the top of the tank, allowing the blades 2a of the turbine 2 to rotate at high speed, thereby mechanically shatter the foam formed and drive the axis 3 of one or more mixers 4. Container 1 has a fluid inlet 17 and a bioprocessing slurry 18 and several sensor insertion points

- 819 shaped so that the sensor can be inserted into the bioprocess broth 6. Figure 2 shows a side view of a disposable bioreactor system with a mixer and mechanical antifoam. Figure 2 shows the vessel 1 of the bioreactor system and the tubes 7, 8 through which air enters the bioreactor vessel and the tube 11 through which the air exits the vessel. Container 1 has a fluid intake location 17 and a bioprocessing slurry collection point 18 and several sensing insertion points 19. The figure shows four sensing insertion points 19, allowing simultaneous monitoring of key parameters in the bioprocessing slurry 6, such as optical density, amount of dissolved oxygen, pH and temperature.

Figure 3 shows top views (position C) showing the container 1 of the bioreactor system, the tube 7 through which air enters the bioreactor tank and the tube 11 through which the air exits the tank, the insertion points of the sensors 19 and the insertion point of liquids 17 into container 1.

Figure 4 shows a cross-section from above in position A of the bioreactor system, in position above turbine 2 with blades 2a, which is secured via the top slider 12 to the inside of the top of the tank

And the bioreactor system. The air that enters the tank through tube 7 drives the blades 2a of turbine 2. The picture shows the insertion points of the sensors 19, the fluid intake site 17 and the bioprocessing slurry extraction point 18.

Figure 5 shows a cross-section from above in position B, in position above mixer 4 and ventilator 5. A tube 8 with a regulator for air intake 10 into the bioprocessing broth 6 is visible.

Figure 6 shows a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam. The disposable bioreactor system consists of a vessel 1, a turbine 2 with blades 2a, which is fixed via the top slide 12 to the inside of the top of the container 1, the axis of the agitator 3, which is attached to the turbine 2, the agitator 4 is attached to the axis 3, and liquid ventilator

5. The pneumatically-driven turbine 2 is driven by a gas stream blown through tubes 7, 8. The tubes have air flow regulators 9, 10 in place, allowing for regulated air intake, thereby providing for regulation of the bioprocess broth ventilation through the aerator 5 and / or rotational speed turbines 2. Rotation of the blades 2a of the turbine 2 simultaneously mechanically breaks the resulting foam resulting from the bioprocess broth 6 and rotates the axis 3 to which one or more mixers are attached 4. The air entering the disposable bioreactor 7, 8 is guided through the wind turbine through the air outlet

II at the top of the tank, allowing the blades 2a of turbine 2 to rotate at high speed, thereby

- 9 mechanically break the foam formed and drive the axis 3 of one or more mixers 4. The picture shows the insertion point of the sensor 19 and the removal site of the bioprocessing slurry 18.

In Fig. 7, as in Fig. 1, a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam is shown, with the turbine 2 being driven by a gas blown into the bioprocess broth 6.

In Fig. 8, as in Fig. 1, a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam is shown, with the turbine 2 being driven by gas blown down the tube 7 into the gas phase 20 of the bioreactor system.

Figure 9 shows, as in Figure 4, a cross-sectional view from above at position A of the bioreactor system, wherein the turbine is driven by a gas blown in several places indicated by 7.

Figure 10 shows, as in Figure 1, a cross-sectional view of a disposable bioreactor system with a mixer and mechanical antifoam consisting of a container 1 with a lid 15, a permeable lid 16 to break the foam formed, and with filters 14 that provide inlet and outlet sterility air.

Figure 11 shows a cross-sectional view of a disposable bioreactor system with a mixer and a mechanical antifoam with a conical shaped top of the container. The disposable bioreactor system consists of a container 1 with a conical shaped top of the container that directs the intake air to the turbine 2, which is fixed via the top slide 12 to the inside of the top of the container 1, the axis of the mixer 3, which is attached to the turbine 2, the mixer 4 is attached to the axis 3, and the fluid aerator 5. The pneumatically driven turbine 2 is driven by a gas flow which is blown through the tubes 7, 8. The tubes have air flow regulators 9, 10 installed, which allows regulated air intake and thus provides for regulation of ventilation. bioprocessing slurries through aerator 5 and / or turbine speed 2. Rotation of the blades 2a of turbine 2 simultaneously mechanically breaks the resulting foam resulting from the bioprocessing slurry 6 through a permeable cover 16 and rotates an axis 3 to which one or more agitators are attached. entering the tubes 7, 8 into the disposable bioreactor is guided through the turbine branch through the air outlet 11 at the top of the tank, allowing the blades 2a of the turbine 2 to rotate with high hit by mechanically breaking the foam formed and driving the axis 3 of one or more mixers 4.

- The bioreactor presented in the invention with a mixer and mechanical antifoam simultaneously solves the problem of efficient material transfer of gas to liquid and the foaming problem resulting from the intensive ventilation of bioprocess broth in disposable bioreactors.

Claims (9)

PATENT APPLICATIONS: 1. A bioreactor system with a turbine attached to the inside of the top of a container, which is driven by a blown gas and which serves to simultaneously break the foam and rotate the axis forming part of the turbine. The bioreactor system according to claim 1, characterized in that the turbine is driven by gas blown into the bioprocessing slurry. Bioreactor system according to claim 1, characterized in that the turbine is driven by a plurality of gas injected into the bioprocessing broth in several places. Bioreactor system according to claim 1, characterized in that the turbine is driven by gas blown into the gas phase of the bioreactor system. The bioreactor system according to claim 1, characterized in that it has a permeable cover for breaking the foam and directing the flow of gas. Bioreactor system according to claim 1, characterized in that the shape of the top of the vessel directing the gas flow to the turbine is conical. Bioreactor system according to claim 1, characterized in that the axis to which one or more agitators is attached. 128. The bioreactor system according to claim 1, characterized in that the gas-tight container has a gas exit point. The bioreactor system according to claim 1, characterized in that the container has a place for withdrawing the bioprocessing slurry. 10. The bioreactor system according to claim 1, characterized in that the container has insertion sites for monitoring optical density of the bioprocess broth and / or sensors for monitoring the oxygen dissolved in the bioprocess broth and / or sensors for monitoring the pH in the bioprocess broth, and / or sensors to monitor the temperature in the bioprocess broth and place for fluid intake.