Patent Application
20140293734
The invention relates to a reactor which is driven in an oscillating-rotating manner about a fixed vertical axis for biotechnological and pharmaceutical applications having process-intensifying properties for mixing, suspension, oxygen transport, heat transfer, irradiation and particle retention, which can be used, without shaft sealing, preferably as a disposable reactor, and thereby ensures a maximum level of process security in terms of cleaning and sterility.
In the highly regulated production of pharmaceuticals, a large expenditure in terms of time, equipment and personnel is apportioned to the provision of cleaned and sterilized bioreactors. In order to avoid cross-contamination reliably in a product change in a multipurpose plant or between two product batches, apart from the cleaning, a very complex cleaning validation is required which may need to be repeated in the event of a process adaptation. This applies not only to upstream processing, USP, that is to say the production of biological products in fermenters, but also to downstream processing, DSP, that is to say purification of the fermentation products. In USP and DSP, use is frequently made of kettles as agitator and reaction systems. Especially in the case of fermentation, an aseptic environment is essential for successful culturing. For the sterilization of batch or fed-batch fermenters, generally the SIP technique is used. In order in the case of continuous process procedure to ensure sufficient long-term sterility, the autoclave technique is also used, which however, requires laborious transport of the reactors to the autoclave and is only usable with comparatively small reactor scales. The risk of contamination during fermentation is particularly critical during sampling and at moving stirrer shafts. The latter are generally equipped with complex sealing systems (e.g.: sliding-ring seals). Technologies which succeed without such penetration of the fermentation casing are preferred because of their greater process robustness.
The downtime of the reactors necessitated by the preparation procedures can be, in particular in the case of short use periods and frequent change of product, of the order of magnitude of reactor availability. The affected steps in the USP of biotechnological production are the process steps of media production and fermentation, and in the DSP, solubilization, freezing, thawing, pH adjustment, precipitation, crystallization, buffer exchange and virus inactivation.
For carrying out the reactions in the USP and DSP, frequently a plurality of reaction conditions must be met simultaneously. For instance, fermentation, for example, in addition to oxygen supply and CO2 removal, requires gentle suspension of the cells, rapid mixing of the media and neutralizing agents for avoidance of overconcentration and also heating of the reaction liquid. Particle retention can also be required, e.g. for the use of perfusion strategies.
In the case of precipitation and crystallization, rapid addition of the precipitant, efficient temperature control and gentle holding of the particles formed in suspension are of particular importance.
Generally, in all process steps of biotechnological production, shallow temperature gradients are required in order not to damage the products. This condition, especially in freezing and thawing processes, leads to considerably increased process times with increasing reactor scale, since no mixing elements can be used in these steps. Heat transport into the reaction medium is limited by the thermal conductivity of the ice layer and also by free convection in the liquid. Long process times, however, can lead to considerable product losses in the presence of proteolytic activity.
Gentle sterilization and virus inactivation of starting materials and product solutions can be achieved by UVC irradiation at a wavelength of 254 nm. The radiation damages the DNA and RNA which lie at the absorption maximum of the viruses and microorganisms and prevents thereby their multiplication, whereas the proteins which are situated in the absorption minimum of the UVC radiation are very substantially retained. A great problem is the depth of penetration of the UVC radiation, which is frequently restricted to only a few tenths of a millimeter in biological media. This makes efficient replacement of the film in the active irradiation zone necessary in order firstly to irradiate all viruses with the required radiation dose and secondly to minimize the radiation load of the products.
The requirement of a constantly renewed boundary layer is also posed in the case of filtration, in order to counteract the development of covering layers which limit the transmembrane flow.
All process engineering steps of mass transport and heat transport, particle separation, UV irradiation and the addition or distribution of solids or additives or gases requires sufficient agitation of the reaction medium. This agitation is ensured, in the pharmaceutical industry, in the stainless steel reactors customarily used there, by means of appropriately dimensioned agitators or by sparging.
Membrane gas introduction is used for gentle oxygen supply of cell cultures. As membranes, gas-permeable silicone tubes are wound onto a cylindrical membrane stator which receive flow from a radially-transporting anchor agitator [WO 2005/111192 A1]. A more than doubling of the exchange area and thereby a significant increase in mass transport can be achieved by paralleling the membrane stators.
Other membrane gas-introduction systems [WO 85/02195 and DE 10 2004 029 709 B4 and DE3428758], in the gas introduction, set up agitators or baskets which are covered with membrane tubes and are moved in a pendulum-like manner in the fermentation solution, or membrane stacks [U.S. Pat. No. 6,708,957 B2], which are swung in the fermentation solution. These membrane gas-introduction systems, however, are distinguished in that they can only be converted to an industrially relevant scale with limitations.
In order to meet the demand for a rapid and flexible charging of the production plant while maintaining maximum cleanliness and sterility, designs for single-use reactors are the subject of constantly growing interest on the market.
Single-use technologies for filtration have long been known. Recently, a single-use technology has also become available on the market for UVC treatment [WO02/038191 WO02/0385502, EP1464342]. Designs for single-use heat exchangers are only available for small scales [EP1464342]. All technologies are operated in continuous flow, so that in addition to a reservoir vessel, the use of pumps and lines is necessary for which cleaning and sanitation plans still need to be provided as before.
There currently exist various commercially available mixing systems which operate on the basis of a plastic bag—single-use technology. These include systems [Hyclone Laboratories, Inc. (http://www.hyclone.com)] which are equipped with blade or magnetic stirrers or circulation pump elements. The systems are available up to a volume of 200 L. [Sartorius AG (http://www.sartorius.de)] offers a single-use system which operates up to a volume of 500 L using a free-floating single-use magnetic stirrer which has no contact with the single-use plastic bag and therefore also no material wear. Single-use mixing systems up to a volume of 10 liters are available at [ATMI, Inc (http://www.atmi-lifesciences.com)]. In this system the material to be mixed is charged into a single-use bag and mixed under rotation. For larger volumes up to 200 L [ATMI, Inc.] offers a single-use bag stirring system which is distinguished in that the stirring element is invaginated into the bag. Mixing in this case is not achieved by a rotary motion around a fixed axis but by a stirring-tilting motion.
In [EP 1 462 155 A1], use is made of a single-use vessel for mixing and dispersing materials by means of a magnetic stirrer which is situated within a protective cage in order to prevent damage to the plastic bag. The product-contact region of the magnetic stirrer unit in this case likewise consists of single-use components.
[EP 1 512 458 A1] demonstrates a solution in which inflatable plastic pillows are integrated in the external or internal region of a single-use bag system. These pillows are alternately pressurized and depressurized again. This induces liquid movements which lead to intensification of mixing and suspension in the vessel.
There are a multiplicity of patents for the use of single-use technology in the fermentation technique sector. In these, in most systems mixing and oxygen supply are achieved via sparging, without further mixing systems being provided [U.S. Pat. No. 5,565,015, WO 98/13469, U.S. Pat. No. 6,432,698 B1, WO 2005/049785 A1, EP 1 602 715 A2, WO 2005/080544 A2]. If a higher oxygen demand is necessary for the culture which cannot be achieved alone via sparging, the sparging can be combined with a dispersing mixing system [WO 2005/104706 A2, WO 2005/108546 A2, WO 2005/118771 A2] or can be overlapped by pumped circulation [WO 2005/067498 A2]. The maximum process volume of a sparged unit is currently up to 1000 liters. In systems having conventional agitators, but which can also be designed as single-use systems [WO 2005/104706 A2, WO 2005/108546 A2], process volumes of up to 10 000 L are achieved.
In the case of sparging, foaming problems can make the use, and the subsequent complex removal, of antifoams necessary in the DSP. The cell stress on bubble rise, in the bursting of the gas bubbles at the surface, and in particular in the foamed destruction, is problematic in cell culture systems, since the cells can be permanently damaged by the resultant high shear forces which are introduced. This applies all the more when sparging is combined with a dispersing agitating system, that is to say an agitating system comminuting the gas bubbles. The damaged cells release proteins, the removal of which can lead to considerable product losses during workup. To maintain acceptable cell vitalities, the oxygen input into the abovementioned bioreactors and therefore also the cell density which is achievable must be restricted. The restricted cell density ultimately reduces the space-time yield of the fermenters and the capacity of the total plant. Since a precondition for reliable upscaling in most cases is not considered technically as met, in the sparged single-use reactors, the volume enlargement must be achieved by complex paralleling of the systems. If the fermenters are operated as proposed using standard agitating systems, although the volume which can be processed increases into the range of the permanently installed plants, the risk of contamination can only be managed with comparable technical expenditure, for example by the use of damped sliding-ring seals. The great technical complexity and expenditure on personnel of such installations, however, largely emphasizes the advantages of the single-use concept.
Other single-use systems provide the necessary gas-introduction rate of the culture by means of membrane or surface gas introduction. In this case the necessary exchange area for gas transport is provided either via a membrane permeable to the gases to be transferred, or via an open boundary area to a gas space. Since no direct gas introduction to the cell culture media proceeds, the particle stress in these reactors may be categorized as low.
[U.S. Pat. No. 5,057,429] describes a system in which an inner semipermeable flat bag which is filled with cell suspension is surrounded by a further bag which is filled with nutrient solution and enriched with oxygen. Nutrient transport and oxygen transport are intensified via a tilting motion of the bags. The maximum process volume of a unit is only a few liters. The oxygen input is considerably restricted by the low oxygen solubility in the charged medium and the comparatively low surface area of the membrane. Compared with standard membrane gas-introduction devices [WO 2005/111192 A1] having specific exchange areas in the order of magnitude of 30 m2/m3 in 100 L reactors, in this arrangement, only a maximum of 10% of this exchange area can be achieved. In both cases, the available exchange area, furthermore, decreases in proportion to the scale enlargement.
Other surface gas-introduction systems likewise operate with a flat bag which is clamped on a shaking apparatus. The bag is only partially filled so that a free surface having a gas space lying thereabove is formed. By means of a seesawing motion or an eccentric rotary motion, the culture medium is mixed, the nutrients which are fed are distributed, cell sedimentation is prevented and the surface is agitated [U.S. Pat. No. 6,190,913 B1, WO 00/66706, U.S. Pat. No. 6,544,788 B2]. In this technology the culture is supplied with oxygen via the free surface. The motion is continuously adapted in such a manner that the flow is gentle and the cells are not exposed to strong shear. The maximum process volume of a unit is currently 580 liters. Although this technology provides a gentle gas-introduction mechanism, it is restricted in conversion to an industrial scale. The height of the bag must be kept approximately constant so that an increase in volume at constant surface area to volume ratio can only be achieved in the two horizontal spatial directions. Upscaling can therefore only be achieved via technically complex paralleling.
The technologies available on the market make use, for freezing, of large stainless steel reactors which are supplied with cooling liquids, or small flat plastic bags which are frozen in a secondary manner via heat-conducting surfaces or by means of convective cold air. In both cases there is no possibility of agitating the product during the freezing process, which considerably lengthens the cooling and freezing processes. The metal vessels are expensive and require large storage areas in the temporary storage. Thawing is lengthy, since the liquid motion between ice block and container wall proceeds only via free convection, comparably to that in freezing. For thawing the plastic bags, these are cut open in the frozen state and subsequently charged into a stirred reactor. The procedure of cutting them open is labor-consuming and contributes to fouling of the working environment. The thawing process is time-intensive, because the ice blocks which float on the surface are hardly reached by the hydrodynamics prevailing in the reactor. Product losses in the course of the long thawing phases are therefore unavoidable.
In the employment of all of the reactors listed here, considerable losses must be accepted in performance and upscalability. In many cases, without sufficient scalability, apart from the lack of performance, an economic benefit cannot be guaranteed. Scaleup here can only be achieved at the cost of increasing complexity and decreasing the economic benefit, such as, for example, by paralleling a plurality of reactors or by the additional use of technically complex solutions (for example sliding-ring seals built into the plastic bags).
A reactor which can be scaled up to the industrial scale of 1 m3-10 m3, guarantees a very high level of sterility comparable to autoclaving by avoiding shaft seals and the problems of cleaning, permits simultaneously intense and gentle liquid motion and can be installed with low expenditure on equipment and personnel, is therefore a clear gap in the currently available range of technologies.
It was an object of the present invention to produce a reactor, in particular for pharmaceutical applications, which, even on large reactor scales, has very good reaction properties for carrying out biological, biochemical and/or chemical reactions with respect to mixing, distribution, suspension, solubilization, mass transport and heat transport, filtration and irradiation, or combinations thereof, and is preferably simple in handling, meets the high requirements of the pharmaceutical industry with respect to cleaning and sterility and contributes to increasing process robustness and to increasing the space-time yield.
The object was achieved by a reactor comprising a reactor vessel and a drive unit, characterized in that the reactor contents, which can be taken up by the reactor vessel, are set in oscillatory-rotary motion about the fixed, preferably vertical, axis of the reactor by the drive unit, wherein the mechanical power input into the reactor contents is enabled by a suitable shell form of the reactor and/or the reactor vessel and/or by internals installed statically in the reactor and/or in the reactor vessel. The reactor is preferably constructed as a single-use reactor.
By means of the internals, distribution processes and/or mixing reactions can be carried out in a simple manner and with the same intensity as in a conventional stirred tank. In this design, a shaft passage can be dispensed with completely. The internals again may be supplied with material streams or energy streams via the side facing away from the product, which streams are introduced into the medium or discharged therefrom by diffusion, convection, heat conduction and/or radiation. In this manner, for the first time, in addition to mixing, numerous process engineering unit operations such as gas distribution, oxygen input by means of membrane gas introduction, heat transport, irradiation and/or particle retention can be carried out in a single-use reactor in a gentle manner and with an efficiency comparable to a stirred tank. The reactions and transport processes proceed in this case directly at the internals. Therefore, the sites of the greatest hydrodynamic energy density and greatest reactivity are identical, or in the case of reactions within the membranes, at least spatially close. No further installations (for example agitators or pumps) are required for transport of the fluids to the reaction site. Since the amount of energy introduced into the liquid is only that which is actually required for carrying out the reaction, these reactions can consequently be carried out in a particularly low-shear manner.
The latter is of critical importance, in particular in the case of shear-sensitive cultures having animal or plant cells which, e.g. during fermentation, must be supplied with oxygen. Because of the high shear forces, here sparging frequently cannot be used, so that generally low-shear membrane gas-introduction is employed. When the static mixing elements in the reactor according to the invention are constructed as tube modules, as described hereinafter, a very high oxygen input or CO2 removal can be ensured using a specific tube or exchange area of more than 30 m2/m3, which is significantly enlarged compared with the prior art, in a single-use reactor without rotating sealing elements, even at large reactor scales.
The reactor has, in particular, a ratio of height to mean diameter of 0.2-2.0, preferably 0.6-1.2 and particularly preferably 0.8-1.0. As a result, tilting moments caused, e.g. by unbalanced masses, can be reduced and a possibility of operation from above is ensured, despite an erection space requirement which can be achieved without problem even on a large scale. In contrast to the slim reactors introduced in biotechnology, such a broad reactor design offers the possibility of dispensing with accommodating reactors in expensive high-rise buildings in favor of erection in cheaper shed-shaped facilities.
Preferably, internals installed in the reactor vessel are provided which provide functionalized surfaces oscillating relative to the drive unit for carrying out physical, biological, biochemical and/or chemical reactions at and/or in membranes. The functionalized surfaces can be provided, in particular, for gas introduction via semipermeable membranes, for gas distribution, for liquid distribution, for irradiation, for filtration, for absorption, for adsorption, for analysis and also for cooling and/or heating.
The invention further relates to a gas-introduction module suitable for such a reactor, in particular a gas distributor or a membrane module which are, in particular, a part of the reactor according to the invention and are described hereinafter by way of example in the installed state. The membrane module which is preferably designed as a tube module has, in particular, essentially vertically arranged permeable, in particular tubular, membranes, through which gases, such as oxygen and carbon dioxide, but no liquid, can pass, in such a manner that oxygen and/or other gases can be introduced into the reactor in a low-shear manner. The membranes can be arranged to be fixed or movable in the reactor and are particularly preferably constructed in such a manner that they can be moved relative to the inert fluid, so that not only gas introduction, gas distribution but additionally also a mixed flow can be induced. In particular, a plurality of groups of membranes or membrane tubes arranged adjacent to one another are provided which provide an exchange surface area required for membrane gas-introduction. For example, the membrane module which is constructed as a tube module and the flat membranes are constructed to be essentially immovable at least relative to the reactor, and only the reactor is driven by the drive unit, in such a manner that the gas-treatment module can be provided as required without expense on construction, in particular as an additional unit which can be connected as desired.
In a preferred embodiment, the membranes are constructed so as to be microporous and permit uniform low-shear distribution of gas bubbles, in particular microbubbles, over the reactor cross section or in the reactor volume without the aid of additional agitator elements. Preferably, the microporous membranes are constructed having core widths of 0.05-500 μm which can be provided via invaginations in the base of the reactors. In this simple manner, bubble coalescence can be successfully prevented. Membranes below 0.5 μm are particularly preferred because particularly fine gas bubbles are generated and an additionally sterile barrier can possibly be dispensed with.
In a preferred embodiment of the gas-introduction module, a first holding profile and a second holding profile are provided between which an elongated, in particular tubular, membrane can be arranged so as to be lead to and fro. The membrane in this case can be arranged in a zigzag shape or a meander shape. As a result, using a single membrane, a particularly large surface area can be provided for gas introduction to the reactor contents.
Preferably, the membrane of the gas-introduction module has a membrane film which is comparatively thin with respect to the total thickness of the membrane. The membrane film is preferably connected in a two-dimensional manner to an open-pore material, such as, for example, foam. In particular, the open-pore material is at least for the most part encased by the at least one membrane film. The open-pore material makes possible uniform convective gas transport through the membrane, in such a manner that essentially the entire membrane film can be charged with gas. Since the membrane film is connected to the open-pore material, the membrane film is simultaneously prevented from being able to inflate at elevated pressures. As a result, a gas-introduction module membrane which is further developed in this manner can be operated without problems even at high pressures, so that with comparatively low material usage, a high volumetric flow rate can be provided for gas introduction to the reactor contents.
In a preferred embodiment, the gas-introduction module is at least in part a part of the reactor vessel of the reactor according to the invention. For this the reactor vessel can have at least two, in particular exactly two, subpieces, while the gas-introduction module has a frame, using which the membranes can be fastened. The subpieces can be connected to the frame, for example by adhesion, in order, together with the frame of the gas-introduction module, to be able to form the reactor vessel of the reactor. For example, two shell-shaped subpieces are provided which, at two end sides facing away from one another of a frame which is shaped essentially rectangularly, can be stuck to the frame. The sides of the frame facing the reactor contents form a part of the shell surface of the reactor vessel. Via the sides of the frame facing away from the reactor contents the membranes of the gas-introduction module can be supplied with gas, for example oxygen, without throughlines needing to be provided therefor, which throughlines would have to be passed through the subpieces.
Preferably, the reactor vessel is lined on an inside at least in part with a permeable membrane for gas introduction to the reactor, in order to improve the gas introduction and avoid dead water zones or laminar boundary layers of the flow. For this, it is, in particular, sufficient merely to form the side of the membrane facing the reactor contents by a membrane film.
Further fields of application for low-shear reactors are precipitation or crystallization of proteins. These operating steps sometimes arise in combination with heat exchange, for example in the plasma fractionation of animal and human blood plasma, and protein purification. In both processes, a particle size distribution which is narrow and shifted to the largest possible particle diameters is required in order to avoid blockage problems and product losses in subsequent particle separation. If the internals are used in whole or in part as distributor layers for the more uniform distribution of the precipitants in the reactor space, over-concentrations may be substantially avoided which contribute to seed formation and thereby to generating very small particles. In addition, the axially transporting, eccentrically arranged, agitating elements in the precipitation reactors which are generally preferred for the macromixing may be avoided, which elements have a particularly intense shearing action on the particles. The reactors, on the product-contact side, are fabricated from materials which are known to those skilled in the art and are stable to solvents and dissolved substances.
Stability to dissolved substances is likewise an important requirement in protein solubilization. The chemicals used therefor in part have the disadvantage of attacking the stainless steel surfaces of standard reactors. The novel reactor concept according to the invention offers the alternative of a broad range of inert materials which is known to those skilled in the art.
A further reaction which can be carried out in the novel mixing reactors is irradiation of the reactor contents for the purpose of sterilization and virus inactivation. Irradiation proceeds within the single-use reactor, e.g. by means of UV irradiators which are positioned in the vessel wall and/or in the built-in elements. Support walls and bags are fabricated from transparent, UV-radiation-transmitting materials which are known to those skilled in the art, the support walls preferably of quartz glass, PMMA or Makrolon, and the bags can be fabricated, depending on the application, e.g., of fluoroelastomers, PMMA or Makrolon. A problem in the UV irradiation of biological media is the frequently extremely limited depth of penetration of the UV rays which, depending on the turbidity, can penetrate only a few tenths of a millimeter of the medium. The good mixing motion and the permanent intensive replacement of the media-side boundary layers means that also the reactor zones far from the boundary layer can be intercepted by the radiation, without the products being impermissibly damaged in the case of too long a residence time in the reactive zones. In this manner, sterilization and inactivation can be carried out for the first time even in single-use large reactors under sterile conditions with large degrees of microbial depletion and small product losses.
Further reactions which can be carried out in this reactor are physical, biological, biochemical and chemical reactions which proceed at least on, and in part in the interior of, functionalized membranes, for instance, for example but not as a restriction, enzymatic reactions, membrane adsorption processes or reactive extractions.
A further reaction which can be carried out by the mixing reactor is freezing and thawing which is required at various points of a biopharmaceutical process, in order to avoid, e.g., a product loss caused by time-dependent decomposition, in the expectation of a release analysis. Using the novel mixing reactor, entire product batches can be frozen, stored in a space-saving manner and thawed out in the same reactor. Freezing and thawing processes proceed in the agitated stage and therefore permit, for intensification and shortening the time of the processes, the use of higher temperature differences between the heating or cooling medium and product solution. Portioning over a plurality of bags and also manual removal of the bags by cutting them open and the fouling of the working environment caused thereby no longer occur.
By means of the process-intensifying internals, the application limits of existing single-use technologies are considerably expanded, in such a manner that the novel reactors can also be employed in scales which are considerably greater than those available hitherto.
The reactor is constructed, in particular, as a single-use reactor which can be discarded after it has been used. For this, the reactor vessel can be produced from a stable, preferably multilayer, polymer material or a polymer material which has been applied to stabilizing grid structures and supporting the intended process engineering unit operations. Preferably, the reactor vessel is connected to a housing which is at least in part adapted to the shell form of the reactor. For this, the reactor vessel which is preferably flexible and/or constructed so as to yield can be inserted and/or suspended into the vessel as a positive fit and/or friction fit. Preferably, the reactor vessel is additionally or alternatively detachably attached to the housing, in particular by reduced pressure. For example, a trough adjacent to the reactor vessel can be provided, to which trough a reduced pressure can be applied in order to fasten the reactor vessel.
Particularly preferably, the vessel and the reactor have at least in part an angular cross section, preferably biangular to octagonal, particularly preferably triangular to quadrangular cross section, and have flat (45), pyramidal (41) or tetrahedral bases.
In this case, the cross sectional shape can also alter in an axial direction over the height of the housing. For instance, the housing can, for example, be constructed in the upper region in a cylindrical or square shape and in a lower region rectangular, square, pyramidal, tetrahedral etc. By a rotary motion of the reactor vessel (46) thus designed, liquid flows (50) can be generated. In addition, the vessel can form internals within an outer wall of the vessel, by which internals the reactor can be accommodated in a non-slip manner, and which simultaneously act in a flow-baffling manner in order to improve the mixing of the reactor contents. The housing can be set in oscillatory-rotary motion about the fixed, preferably vertical, axis of the reactor by the drive unit, in such a manner that direct coupling of the drive unit to the reactor vessel itself is not required. As a result, most components can be reused, so that only the, if appropriate, specially shaped single-use reactor need be disposed of, which in principle does not require any additional mixing elements in order to achieve a low-shear mixing. Preferably, the housing is rotatably mounted, in particular suspended, so as to be movable in an essentially vertical direction. The housing, as a result, can, for example, be inserted simply into a holder or an axial bearing from the top using a crane or from the bottom by means of a lifting platform, in such a manner that for various housing or reactor vessel types the same drive unit and/or the same measurement technique can be employed.
Preferably, the reactor is positively coupled to the drive unit in such a manner that the acceleration and braking of the reactor rotation proceeds with an essentially constant angular acceleration or deceleration. As a result, the speed of rotation of the reactor changes linearly with time in each movement phase of the rotary oscillation. Control modules connected intermediately are not required in this simple reactor movement so that, for example, according to a preferred embodiment for implementation of the oscillatory reactor motion, a pendulum drive can be used. As a result, e.g. the release of electromagnetic rays which can cause, e.g., faults of sensors, can be drastically reduced. In particular, as a result of the constant angular acceleration in each phase of the oscillatory-rotary reactor motion, instantaneous peak values of the hydrodynamic shear forces on suspended particles (e.g. animal cells) are kept comparatively smaller than in other forms of motion of the reactor.
It has been discovered, surprisingly, that a comparatively small angular amplitude is sufficient for the oscillatory-rotary motion of the reactor in order to achieve good mixing and/or sufficient intensification of transport processes. In particular, it is hardly necessary to implement 3600° rotations, that corresponds to 10 rotations, of the reactor, so that structurally complex solutions for connecting the oscillatory-rotary reactor to the static surrounds (e.g. for the feed and removal of media and gases, of electrical energy and electric signals) are hardly required. The reactor can carry out an oscillatory-rotary motion in which the angular amplitude a is in the range 2°≦|α⊕≦3600°, preferably 20°≦|α|≦180°, particularly preferably 45°≦|α|≦90°. In particular, approximately |α|=45° or |α|=90°, wherein deviations of ±5° can be present. In total, therefore, the oscillatory motion sweeps an angle of 2|α|.
Experiments have found that when the power input is increased, states of motion can be established in this reactor in which gas bubbles are introduced into the reactor. For the cells which are not damaged by sparging, a very simple gas distribution can be achieved in this manner in a preferred polygonal, particularly preferably 2-4-angular embodiment of the reactor according to the invention without cost-intensive internals. Surprisingly, it has been found that unwanted foam development first, as expected, increases with increasing reactor agitation, in order, however, then after exceeding a maximum foam height to decrease again to easily manageable foam heights of a few centimeters. The cause of this highly astonishing phenomenon of this foam destruction is that in these states of motion of the liquid, not only the gas which is situated in the headspace, but also the foam itself is drawn in by the surface. The foam, by being sucked back below the liquid surface, is redissolved gently without application of shear forces, that is to say with strict avoidance of bursting of gas bubbles. In particular, a wave flow can be established by which a part of the reactor contents which are situated on the surface are transported into the interior of the reactor contents. In this preferred reactor type, therefore foam formation can be substantially suppressed and simultaneously particularly gentle and effective surface gas-introduction can be achieved. The use of the oscillating foam destroyer, however, is in no way restricted to reactors with surface gas-introduction, but, according to a particularly preferred embodiment, may advantageously be used generally in sparged reactors. Therefore, preferably, in the reactor according to the invention, in particular the intensity of the oscillatory-rotary motion can be set in such a manner that, at the surface of the reactor contents, a wave flow can be generated which transports a part of the reactor contents which are situated at the surface into the interior of the reactor contents.
In a preferred embodiment, the reactor vessel has an elongate fluorescence sensor running essentially in the peripheral direction to the axis of the reactor, using which, in particular a pH and/or an oxygen concentration of the reactor contents can be detected. For contact-free detection, an optical detection apparatus at a distance from the reactor vessel is provided, which gives off, for example, a light flash, in order to be able to determine, from the reaction of the fluorescence sensor to the light flash, the desired measured value. In particular, the detection rates and the oscillatory-rotary motion are selected in such a manner that the fluorescence sensor is optically detected at various part-surfaces. It is therefore possible to irradiate the fluorescence sensor at different points, so that bleaching of the fluorescence sensor by “photo bleaching” is prevented and the service life is significantly increased.
The invention further relates to a sparged reactor having a reactor vessel which has a polygonal cross section at least in the region of a liquid surface of reactor contents taken up by the reactor vessel, which reactor contents are charged with gas bubbles via the surface or porous membranes and, for the purpose of foam destruction, are set into an oscillatory-rotary motion such that foam on the surface of the reactor contents is transported into the interior of the reactor contents. The sparged reactor can, in particular, be formed and further developed as described above. The sparged reactor, therefore, is constructed in such a manner that, additionally, or alternatively, it can be a foam destroyer.
Preferably, a process is provided in which a reactor or sparged reactor is used which can be formed and further developed as described above. The reactor is particularly preferably used for suspending bioreactive substances. Therefore biological materials such as, for example, animal and/or plant cells and/or microorganisms, can be provided which are intended to be suspended in a liquid substrate in order, in particular with continuous addition of oxygen, to chemically react substances present in the substrate. The oscillatory-rotary motion of the reactor and the power input can, in particular, be set in such a manner that foam formation on the surface of the reactor contents is minimized. It is already sufficient for this for the oscillatory-rotary motion of the reactor to be at a comparatively small angular amplitude |α| of ≦3600°, clockwise and anticlockwise. The reactor or the sparged reactor is used, in particular, for the preferably low-shear destruction of foam which can form on mixing and/or gas introduction. The destruction of the foam proceeds, in particular, by solubilizing the foam, which can be drawn into the interior of the reactor contents by the flow induced in the reactor vessel. That is to say the foam which is drawn in can collapse in a low-shear manner in the interior of the reactor contents.
The invention will be described in more detail hereinafter with reference to the accompanying drawings referring to preferred examples, to which the invention is not restricted.
The drawings show the following:
a: a schematic simplified side view of an installed reactor,
b: a schematic perspective view of the reactor of
a: a schematic diagram having rotary oscillation suitable for the reactor,
b: a schematic diagram for comparison of the O2 introduction in various gas-introduction methods,
a: a schematic side view of the reactor of
b: a schematic plan view of the reactor of
c: a schematic sectional detail view of the reactor of
a: a schematic simplified side view of the installed reactor in a further embodiment,
b: a schematic perspective view of the reactor of
a: a schematic perspective view of the reactor in a further embodiment,
b: a schematic perspective view of the reactor in a further embodiment,
c: a schematic perspective plan view of the reactor of
d: a schematic sectional detail view of the reactor of
e: a schematic plan view of the reactor of
f: a schematic sectional view of the reactor of
g: a schematic plan view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment,
b: a schematic plan view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment,
b: a schematic sectional view of a silicone tube suitable for the reactor,
c: a schematic sectional view of a module having the silicone tube of
d: a schematic sectional view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment,
b: a schematic sectional view of the reactor of
c: a schematic plan view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment,
b: a schematic sectional view of the reactor of
c: a schematic plan view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment in a first state,
b: a schematic sectional view of the reactor of
c: a schematic sectional view of the reactor of
d: a schematic sectional view of the reactor of
e: a schematic sectional view of the reactor of
f: a schematic sectional view of the reactor of
a: a schematic sectional view of the reactor in the installed state in a further embodiment,
b: a schematic sectional detail view of the reactor of
c: a schematic sectional detail view of the reactor of
a: a schematic sectional view and a schematic plan view of the reactor in the installed state in a further embodiment,
b: a schematic sectional view and a schematic plan view of the reactor in the installed state in a further embodiment.
a: a schematic perspective exploded view of the reactor in a further embodiment,
b: a schematic sectional view of the reactor of
c: a schematic plan view of the reactor of
a: a schematic perspective exploded view of the reactor of
b: a schematic sectional view of the reactor of
c: a schematic plan view of the reactor of
d: a schematic perspective view of the reactor of
e: a schematic perspective view of the reactor of
a: a schematic sectional view of a membrane suitable for the gas-introduction module,
b: a schematic sectional view of a membrane suitable for the gas-introduction module, in a further embodiment,
c: a schematic sectional detail view of the membrane of
d: a schematic sectional view of a membrane suitable for the gas-introduction module, in a further embodiment,
e: a schematic sectional plan view of the membrane of
a: a schematic sectional view of the reactor in a further embodiment,
b: a schematic plan view of the reactor of
c: a schematic perspective view of a part of the gas-introduction module for the reactor of
a and 1b show a reactor vessel, designated reactor 5, of the reactor according to the invention having a drive unit without process-intensifying internals. The medium 4, a substrate or buffer solution, a fermentation solution or a product solution, is contained in the reactor 5 which, in the case of the particularly preferred use as single-use reactor for improving stability, is produced from stable, preferably multilayer, plastic films which are known to those skilled in the art. The mass of the filled reactor 5 which, for strength reasons, can only be burdened with limited tensile and shearing forces, is taken up in the vertical direction from the base 20 of a surrounding vessel and in the side direction via the shell 6 thereof. For simple installation of the reactor 5, the shell 6 can be opened via a door 7. During the process the base 20 which is rotatably mounted on the bearing 8 is set in oscillating rotation 15 via the drive table 10. The position of the drive axle is preferably fixed in order to avoid transverse forces caused by eccentricity on the reactor 5, or the system consisting of shell 6, base 20 and drive table 10. Transverse forces pose considerable problems on upscaling. The angle of the drive axle can be chosen, in principle, as desired between 0 and 90° to the horizontal. Angles around 90° to the horizontal are among the particularly preferred embodiments, because as a result a comparatively simple bearing mounting of the reactor and the drive unit is possible. In this type of bearing, the head region of the reactor 5 remains substantially unloaded and permits a simple access to the reactor interior by connection lines and sensors. By means of a simply achieved size adaptation of shell 6 and reactor 5, smaller reactors can also be operated on the same base 20, which increases the flexibility of production, in particular in the case of frequent product changes.
a shows suitable rotary oscillations 15, e.g. having a rectangular 158, linear 157 or sinusoidal 156 course of angular velocity with time. The period 160 and amplitude 162 of the rotary oscillation 15 depend on the geometry and size of the reactor 5 and internals thereof and the desired mechanical power input which is required to carry out the process step. A low-shear motion can be induced when losses due to flow around the internal elements and thereby relative velocity between the internal elements and the fluid can be kept as constant as possible. In order to achieve this, the fluid is expediently first accelerated into the one direction with a sinusoidal velocity impulse 156 of the internal element and later decelerated in order, on passing through zero of the rotary velocity, ultimately to be accelerated and decelerated in the opposite direction. If the torque of the drive used permits high angular accelerations of the reactor, rectangular impulses 158 may be implemented to an approximation. However, these lead to a considerably greater range of velocity distribution in the reactor and thereby, with comparable mechanical power input, to an increase of the shear stress of suspended particles. This drive mode will generally be avoided on the culture of shear-sensitive animal cells. In the suspension of sedimented particles or on mixing in additives, these additional mixing effects, in contrast, are absolutely desired.
In the case of membrane gas introduction, high specific exchange areas of significantly more than 30 m2/m3 can be ensured in a single-use reactor without rotating sealing elements and, what is more, in very large reactor scales.
In the case of generation of coarse or fine gas bubbles via membranes, use can be made of, for example, the oscillating motion of microporous invaginations 150, in order to distribute gas bubbles in the liquid 11 uniformly over the reactor cross section.
a shows by way of example, but not restricting the invention, in what manner the base 20 rotatably mounted on the bearing 8 can be driven via a gear wheel 12 using an electric drive 14 which is installed in the drive table 10. Alternative drive possibilities to electric drives 14 could be provided via magnetic forces, induction forces, pneumatics or hydraulics. For heating/cooling the medium 4, the base 20 can be equipped with a cavity 32 in which an electric (for example a heating mat) or a heat exchanger 18 through which a heating/cooling medium flows can be accommodated. To improve the heat transfer it is advisable to fill the cavity 32 with a readily heat-conducting heat transfer medium, e.g. water or oil. The heat exchanger is supplied via a central line 30 which is connected via tubes or cables to the energy supply, i.e. to a heating/cooling circuit or to electricity. Addition or withdrawal to or from the reactor 5 can be performed via central 27 or offcenter 24, 28, passages through the head of the reactor 5. Using the lance 28, addition also into the depth in the reactor 5 can proceed. In the case of offcenter addition, the lance 28 acts as a flow resistance to the surrounding medium 4, so that at the introduction site, corresponding to the chosen intensity of the rotary oscillation 15, a liquid flow favoring mixing can be produced. The passageways 24, 26 and 28 are likewise suitable for bringing commercially available sampling systems and sensors for measuring temperature, gas content, ion concentration, optical properties, particle concentration and cell vitality into contact with the medium 4 or the gas space for the purpose of process control. The introduction of thermally or chemically presterilized and calibrated systems can proceed at the start of the process under a safety cabinet. The sensors are customarily fastened to the port using a screw connection and sealed to the inner flanks of the passageways by means of an O-ring. Preference is further given to sensors based on fluorescent reactive dyes which are simply applied to the reactor wall in order to interact with the medium. Excitation and measurement of the layers can proceed non-invasively from the outside, which eliminates the risk to sterility of sensor introduction. The load-bearing capacity of reactors fabricated with plastic can be increased in the region of the passageways by means of welded or glued reinforcements 25 (see also
An expedient embodiment of the single-use reactor which does not restrict the invention is shown in
A particularly simple, and nevertheless effective, method for transmitting the rotary oscillation 15 from the reactor walls to the medium 4 can proceed without flow internals, just by the choice of a suitable reactor geometry. If, as is shown in
In the illustrative example shown in
A considerable reinforcement of the mixing motion is possible using internals.
a to c show by way of example of the cylindrical reactor 5, which is not restricting to the invention, that, for improving the oxygen introduction, tube modules 72 can relatively simply be accommodated in a reactor 5 as in the mixing appliance shown in
a to 8c show an alternative reactor design which is particularly advantageous compared with the mixing reactor in
In
Using the arrangement in
In
In
In
In
In the illustrative example of the reactor shown in
In the illustrative example of the reactor shown in
In the alternative embodiment of the membrane 74 shown in
In the embodiment of the reactor shown in
The reactor shown in
By means of the process-intensifying internals and thereby the physical, biological, biochemical and chemical reactions which can be carried out in and on membranes, e.g. for gas introduction, gas distribution, liquid distribution, particle retention, irradiation and/or heat feed and removal, the application limits of existing single-use technologies are considerably extended, so that the novel reactors can also be employed on considerably greater scales than those previously available.
| Number | Date | Country | Kind |
|---|---|---|---|
| 10 2006 018 824.1 | Apr 2006 | DE | national |
This is a Division of application Ser. No. 14/047,531 filed Oct. 7, 2013, which is a Division of application Ser. No. 12/297,987 filed Nov. 26, 2008, now U.S. Pat. No. 8,602,636, which is a 371 of PCT/EP2007/003521 filed Apr. 23, 2007, claiming priority of German Applicaton DE 10 2006 018 824.1 filed Apr. 22, 2006, the disclosures of which are all incorporated herein by reference.
| Number | Date | Country | |
|---|---|---|---|
| Parent | 14047531 | Oct 2013 | US |
| Child | 14278527 | US | |
| Parent | 12297987 | Nov 2008 | US |
| Child | 14047531 | US |
