Disposable Bioreactor Systems and Related Methods

Abstract

A method for enhancing mixing and aeration of a liquid reaction medium in a toroidal bioreactor vessel includes dispensing a liquid reaction medium into an interior of a toroidal bioreactor vessel, the interior being bounded by an inner surface, a textured surface being arranged on at least a portion of the inner surface, the textured surface having a plurality of upstanding protuberances. The toroidal bioreactor vessel is rotated in an orbital motion such that there is a resonant frequency traveling wave of the fluid orbiting in one direction in the interior of the toroidal bioreactor vessel when a particular orbital speed is imparted to the toroidal bioreactor vessel.

Claims

1. An apparatus comprising: (a) a hollow toroidal bioreactor vessel having an interior bounded by an inner surface, a textured surface being arranged on at least a portion of the inner surface, the textured surface having a sinusoidal shape pattern and being comprised of a plurality of upstanding protuberances; and (b) a drive including a motor driven platform, wherein when the toroidal bioreactor vessel rests on the motor driven platform and a fluid is disposed within the interior of the toroidal bioreactor vessel, operation of the drive moves the platform orbitally such that there is a resonant frequency traveling wave of the fluid orbiting in one direction in the interior of the toroidal bioreactor vessel when a particular orbital speed is imparted to the toroidal bioreactor vessel, and wherein the orbiting direction of the traveling wave of the fluid is parallel to a direction of periodic oscillation of the sinusoidal shape pattern of the textured surface.

2. The apparatus in accordance with claim 1, wherein the textured surface is located on a bottom of the toroidal bioreactor vessel.

3. The apparatus in accordance with claim 1, wherein the textured surface is located on a side-wall of the toroidal bioreactor vessel.

4. The apparatus in accordance with claim 1, wherein the textured surface is located on both a bottom of the toroidal bioreactor vessel and a side-wall of the toroidal bioreactor vessel.

5. The apparatus in accordance with claim 1, the toroidal bioreactor vessel further comprising a plurality of tubes coupled to at least a portion of the inner surface of the toroidal bioreactor vessel, wherein the plurality of tubes are configured for sparging gas into said toroidal bioreactor vessel.

6. The apparatus in accordance with claim 5, wherein the plurality of tubes is coupled to a bottom or a top of the toroidal bioreactor vessel.

7. The apparatus in accordance with claim 5, wherein the plurality of tubes is coupled to a side-wall of the hollow toroidal bioreactor vessel.

8. The apparatus in accordance with claim 5, wherein at least some of the plurality of tubes extend through the plurality of upstanding protuberances of the textured surface.

9. The apparatus in accordance with claim 1, wherein operation of the drive moves the platform both orbitally and in a rocking motion.

10. The apparatus in accordance with claim 1, wherein when in operation, the drive is configured to move the toroidal bioreactor vessel in a reciprocating motion to cause the fluid to orbit in a reversed direction.

11. An apparatus comprising: (a) a hollow bioreactor vessel comprising: an interior bounded by an inner surface; a textured surface arranged on the inner surface, the textured surface comprising a plurality of upstanding protuberances; and a plurality of tubes coupled to the inner surface and extending through the plurality of upstanding protuberances of the textured surface, the plurality of tubes being configured for sparging a gas into the interior of the bioreactor vessel; and (b) a drive including a motor driven platform, the bioreactor vessel being disposed on the motor driven platform, the drive being configured to move the platform orbitally.

12. The apparatus in accordance with claim 11, wherein when a fluid is disposed within the interior of the bioreactor vessel and the drive is activated, the drive moves the bioreactor vessel such that there is a resonant frequency traveling wave of the fluid orbiting in one direction in the interior of the bioreactor vessel when a particular orbital speed is imparted to the bioreactor vessel, and wherein the textured surface, the plurality of tubes, and the traveling wave of fluid are configured to enhance aeration and mixing efficiency of the fluid.

13. The apparatus in accordance with claim 11, wherein the textured surface is located on a bottom of the bioreactor vessel.

14. The apparatus in accordance with claim 11, wherein said bioreactor vessel comprises a hollow toroidal bioreactor vessel and includes at least one exit port for removal of gaseous by-products.

15. The apparatus in accordance with claim 11, further comprising a plurality of input ports for inserting sensors or process feedstock inputs into the bioreactor vessel.

16. A method for enhancing mixing and aeration of a liquid reaction medium in a toroidal bioreactor vessel, the method comprising: dispensing a liquid reaction medium into an interior of a toroidal bioreactor vessel, the interior being bounded by an inner surface, a textured surface being arranged on at least a portion of the inner surface, the textured surface comprises a plurality of upstanding protuberances; and rotating the toroidal bioreactor vessel in an orbital motion such that there is a resonant frequency traveling wave of the fluid orbiting in one direction in the interior of the toroidal bioreactor vessel when a particular orbital speed is imparted to the toroidal bioreactor vessel.

17. The method in accordance with claim 16, wherein the orbiting direction of the traveling wave of the fluid is parallel to a direction of periodic oscillation of the sinusoidal shape pattern of the textured surface.

18. The method in accordance with claim 16, the method further comprising sparging the liquid reaction medium in the toroidal bioreactor vessel during the rotating step by introducing a gas into the interior of the toroidal bioreactor vessel through a plurality of orifices in the inner surface of the toroidal bioreactor vessel.

19. The method in accordance with claim 18, wherein the toroidal bioreactor vessel further comprises a plurality of tubes coupled to the plurality of orifices in the inner surface and extending through the plurality of upstanding protuberances of the textured surface, the plurality of tubes sparging the gas into the toroidal bioreactor vessel.

20. The method in accordance with claim 16, the method further comprising, prior to the rotating step: determining a resonant frequency for the resonant frequency traveling wave of fluid from dimensions of the toroidal bioreactor vessel; and determining a particular orbital speed for the hollow toroidal bioreactor vessel from the resonant frequency such that the resonant frequency traveling wave of fluid reproduces itself in a single orbital rotation of the hollow toroidal bioreactor vessel during the rotating step.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0016] FIG. 1 shows the prior art type of disposable bioreactor system (e.g., HyClone/Thermo-Fisher) that utilizes a disposable polymeric liner and provides structural support for the liner.

[0017] FIG. 2 shows a prior art pillow disposable bioreactor bag of flexible polymeric material.

[0018] FIG. 3a shows a three-dimensional view of a bioreactor in accordance with the present invention having vertical side walls and which is toroidal in shape.

[0019] FIG. 3b shows two-dimensional views of a toroidal bioreactor in accordance with the present invention as depicted in FIG. 3a and shows alternative embodiments of the vessel having either a circular (12) or rectangular (11) cross-section.

[0020] FIG. 3c shows how the bioreactor of the present invention achieves its enhanced mixing via orbital motion (in this case moving counterclockwise indicated as 16) which creates a travelling wave of the growth medium present in the bioreactor vessel (17).

[0021] FIG. 4 shows a view of the walls of the preferred embodiment of the bioreactor vessel of FIGS. 3a and 3b where sparging is accomplished through the walls of the vessel.

[0022] The walls are shown as corrugated in order to enhance mixing.

[0023] FIG. 5 shows a view of an alternative configuration for a bioreactor vessel in accordance with the present invention where the bioreactor is toroidal but oval in cross-section rather than circular. Again, the vessel can have a cross-section that is either substantially rectangular (5.2) or circular (5.3).

[0024] FIG. 6 shows an embodiment of the present invention which provides for multi-directional motion (i.e., both orbital and rocking) of the bioreactor vessel of the present invention shown in FIG. 3a, with the X and Y axes shown.

DESCRIPTION OF THE INVENTION

[0025] The most desirable single-use bioreactor solution would be one that effectively meets criteria 1 through 3, as set forth above, while at the same time requiring the fewest moving parts. Minimizing the number of moving parts will also tend to yield the least expensive and most reliable overall solution. A novel design for a polymeric bioreactor vessel in accordance with the present invention which uses a minimal number of parts is shown in FIGS. 3 through 6.

[0026] In FIG. 3a, a cylindrical bioreactor vessel 1 having a substantially rectangular cross-section is shown having a hollow core 6, so that the bioreactor is a toroid. Although the design shown in FIG. 3a is shown as having substantially vertical walls, this is not an essential aspect of the invention and a toroidal (donut) shaped vessel not having vertical surfaces is also suitable, as shown in FIG. 3b in cross section as 12. Alternatively, only one wall (either inner or outer) may be vertical and the other curved.

[0027] The vessel can suitably be of a rigid biocompatible polymer. Alternatively, the vessel can be fabricated in whole or in part of a flexible (non-rigid) polymer since the hydrostatic pressure of the liquid reaction (growth) medium present within the vessel together with the pressure of the sparging oxygen or oxygen containing gas will enable even a bag made of non-rigid polymer to substantially retain its annular shape. Suitable rigid polymers include, but are not limited to, USP Class VI approved polycarbonate and polystyrene. Suitable flexible polymers include, but are not limited to, low density polyethylene and ethylene/vinyl acetate copolymer.

[0028] In FIG. 3a, element 2 is a platform equipped with an electric motor or other drive means (not shown) which moves in an orbital motion as indicated by arrow 3 and as further shown in FIG. 3c. This orbital motion, combined with the toroidal shape of the bioreactor, sets up a traveling wave in the bioreactor growth medium. This traveling wave provides both effective mixing and oxygenation of the contents. It should be noted that in the case of a toroidal vessel whether having vertical or curved walls, the liquid motion will follow a pure traveling wave. In both cases, the exact liquid motion can be modeled using computational fluid dynamics.

[0029] Additionally, single-use bioreactor vessel 1 can suitably be equipped with one or a plurality of input ports (one shown as 4 in FIG. 3a) for process feedstock inputs (e.g.: 10 pH buffers, glucose etc.), and/or also for pre-inserted and pre-calibrated sensors (e.g., to measure dissolved oxygen, pH, dissolved CO.sub.2 and the like). These sensors can be either traditional electrochemical sensors and/or disposable and pre-calibrated optical sensors. It is generally more efficacious to have the sensors pre-calibrated in order to avoid breaking the sterile barrier on the bioreactor. Optical probes are available that can be both pre-calibrated and can be gamma irradiated while mounted in the bioreactor. The ability to use gamma radiation allows the pre-calibrated sensors to be inserted in the bioreactor and the entire assembly subsequently sterilized. Port 5 denotes the exit port for removal of gaseous reaction by-products. In FIG. 3a 7 denotes a light source such as a bank of visible or UV light emitting LEDs that can be utilized to promote the growth of plant or algae culture or transiently activate certain genes for multi-drug production from a single cell platform. The lighting is shown on the top, but this is not a necessity; the illumination can additionally or alternatively come from the sides and/or even the bottom of the vessel provided only that the growth media receives enough illumination to enhance photosynthesis or gene activation depending on the purpose of the growth run and the cell being grown. Ideally the LEDs will have a spectrum that is matched to the absorption spectrum of the species being grown. LEDs are now available from the ultra-violet through the near infrared (e.g.: http://www.marktechopto.com/) so matching to the absorption spectrum of the species under study can be easily done. Additionally, the surface of the bioreactor on which the LED's are mounted needs to be sufficiently transmitting in this region to allow the light to pass through to the interior of the bioreactor.

[0030] The bioreactor vessel walls (outer and optional inner surfaces) define a structure that will have a resonant frequency which is determined by the particular configuration (size and shape) of the bioreactor. The resonant frequency of the bioreactor can be readily calculated knowing that the traveling wave must reproduce itself in phase every round trip (Hydrodynamics, Horace Lamb and Russ Caflisch, First Cambridge University Press, 1997). Once this frequency has been determined the bioreactor can be rotated with this circular frequency to thereby set-up resonant traveling wave motion of the fluid (growth medium) inside. The wave amplitude will be chosen depending on the level of mixing and agitation needed.

[0031] In FIG. 3b the top view, 10, of the bioreactor is shown along with the square or alternatively circular cross sections 11 and 12 respectively.

[0032] It is possible to further enhance the mixing and sparging efficiency of the bioreactor vessel of the present invention shown in FIG. 4 by providing a textured surface on the inner floor (bottom) and/or wall surfaces of the vessel (i.e., a surface which is in contact with the growth medium). This texturing can be achieved using baffles, ridges, bumps and/or other upstanding protuberances as shown in FIG. 4, which are preferably pre-molded on all or a part of the inner surface of the vessel. The sparging of the reaction medium is preferably accomplished in this design by bringing the oxygen or air into the bioreactor through a multitude of orifices in the floor and/or walls of the vessel. These orifices can be in the protuberances in the texturing or separate from them. In FIG. 4, the flow direction of the fluid in the annular vessel is indicated as 4.1, (which corresponds to arrow 3 in FIG. 4), the textured surface on the inner wall and/or floor surface of the vessel (the textured surface being shown here for simplicity to have an approximately sinusoidal shape pattern) is indicated as 4.2, and the sparge orifices are indicated as 4.3.

[0033] The optimal patterning (e.g., size, shape and frequency) will be a function of the size of the reactor, the velocity, viscosity, and nature of cell platform and its associated optimized growth medium. The particular patterning which provides optimal agitation can be determined through finite element analysis studies (www.fluent.com) or through empirical experiment. These studies generally include mixing studies as a function of time or number of agitation cycles. Additional computational studies that employ Henry's law (p. 384, General Chemistry, 2.sup.nd Edition, Donald A. McQuarrie and Peter A. Rock, W. H Freeman and Company, New York, 1987) to model oxygen transfer or calculate the oxygen transfer rate are possible. These studies require the finite element analysis code to take into account the surface area of the bubbles created during sparging. For example, a higher number of bubbles having decreased size will increase the surface area available for oxygen transfer.

[0034] In FIG. 4, the sparging gas (normally air or oxygen) is suitably brought to the bioreactor using tubing through the sparge orifices 4.3 which preferably will have a fluid diode or hydrophobic filter at or near its end that allows the sparging gas to flow into the bioreactor, but prevents the liquid (reaction medium) from draining out of the bioreactor vessel through said tubing. The aforementioned texturing of the floor and/or walls therefore does not preclude bringing the sparge gas into the vessel through the floor, top, and/or walls of the vessel. This type of sparging, combined with the traveling wave motion induced by the orbital movement of the vessel, ensures that the cells or microbes in the bioreactor have access to sufficient feed and oxygen to achieve optimal total cell density and/or maximize product yield.

[0035] It should also be noted that the motion in direction FIG. 3a is only shown as orbital, but can also involve rocking in either the X or Y axis (or both) and thereby be a two-dimensional or three-dimensional motion, which can be readily tailored to work in conjunction with the bioreactor dimensions. Such multi-directional motion is shown in FIG. 6 where 6.1 and 6.2 are the plane of the platform when rocked about axis 6.3. The platform holding the bioreactor can also be orbited in one direction according to 6.4, or agitated in both directions as per 6.5. This motion can also be a reciprocating motion where the direction of rotation of the motor which causes the vessel to orbit is periodically reversed. This periodic reversal in the direction of the vessel's orbit can be used to cause more vigorous agitation if/when required during the growth process.