Online concentration system, control method and chromatographic system matched with online concentration system

Abstract

The invention provides an online concentration system, a control method and a chromatographic system matched with the online concentration system. The online concentration system comprises a first input pipeline and a second input pipeline, a first feeding pump and a second feeding pump are respectively arranged on the first input pipeline and the second input pipeline, the output ends of the first input pipeline and the second input pipeline are connected with the inlet end of a tangential flow filter through pipelines, and the intercepting end of the tangential flow filter is connected with a first output pipeline; a first discharging pump is arranged on the first output pipeline; a circulating pipeline is arranged between the inlet end and the intercepting end of the tangential flow filter, and a reflux pump is arranged on the circulating pipeline; the permeation end of the tangential flow filter is connected with a permeate pipeline and a chromatographic system matched with the chromatographic column for use. The device has the advantages that the tangential flow can be increased, the transmembrane pressure is not increased, and the yield is improved.

Description

Technical Field

[0001] The present invention relates to the concentration, separation and purification of downstream biopharmaceutical products, and in particular to an online concentration system, a control method and a matching chromatography system. Background Art

[0002] Chromatographic technology is widely used in many fields such as biology, medicine, and chemistry. Before the product is loaded onto the sample, the pH and concentration of the product need to be controlled. Only after the loading conditions are met can the subsequent separation and purification steps be carried out. Therefore, the current existing processes all involve concentrating and replacing the product, collecting it in a circulation tank, and then loading it onto a chromatographic column for separation and purification. Common methods for concentrating and replacing products mostly use a single pump mode, that is, the bottom outlet of the circulation tank is connected to the inlet of the tangential flow filter through a feed pump, the reflux end of the tangential flow filter is connected to the top inlet of the circulation tank, and the permeate end of the tangential flow filter is connected to the permeate pipe. During use, the sample to be processed is filtered through the tangential flow filter and a buffer solution is added to the retained species to achieve the purpose of concentration and buffer replacement, so that the pH and concentration of the sample to be processed meet the conditions for chromatographic loading. Therefore, it is divided into two separate process steps. Its disadvantages are: (1) the tangential flow filtration of a single pump has low efficiency, takes a long time, and requires more filter membranes; (2) the tangential flow of a single pump requires a circulation tank, and the circulation requires a weight sensor, a stirrer, and a nozzle; (3) two separate process steps require two separate process equipment, which not only takes a lot of time to concentrate and replace the sample before chromatographic loading, but also requires two sets of equipment, and space for equipment use is required.

[0003] Displacement chromatography is performed by connecting a chromatographic column to a direct tangential flow system, without using the recirculation structure of the tangential flow system. However, this method suffers from the inability to coordinate the flow rates required by the tangential flow system and the column. Therefore, it is technically difficult to achieve a direct connection between a tangential flow system and a chromatographic column without using a recirculation structure.

[0004] When the outlet flow rate decreases and the fluidity is insufficient, the product to be concentrated will accumulate on the membrane surface, resulting in a decrease in yield. Summary of the Invention

[0005] To overcome the shortcomings of the prior art, the present invention provides an online concentration system, control method, and supporting chromatography system, which increase tangential flow without increasing transmembrane pressure, thereby improving yield. These systems eliminate the need for a circulation tank, weight sensor, stirring device, or spray nozzle, thus reducing equipment footprint and costs. They can meet the flow rate requirements of the chromatography system and enable online sample loading.

[0006] In response to the above technical problems, the first aspect disclosed in the present invention proposes an online concentration system, comprising a first input pipeline and a second input pipeline; the first input pipeline and the second input pipeline are respectively provided with a first feed pump and a second feed pump, the output ends of the first input pipeline and the second input pipeline are connected to the inlet end of the tangential flow filter through pipelines, the intercepting end of the tangential flow filter is connected to the first output pipeline, and the first output pipeline is provided with a first discharge pump; a circulation pipeline is provided between the inlet end and the intercepting end of the tangential flow filter, and the circulation pipeline is provided with a reflux pump; the permeate end of the tangential flow filter is connected to the permeate pipeline.

[0007] In some embodiments, a mixer is provided between the output ends of the first input pipe and the second input pipe and the inlet end of the tangential flow filter, and the outlet of the circulation pipe is located at the inlet end of the mixer.

[0008] In some embodiments, flow sensors are respectively provided on the first input pipe and / or the second input pipe and / or the first output pipe and / or the circulation pipe.

[0009] In some embodiments, the flow rate ratio between the first input conduit and the second input conduit is used to control the concentration ratio of the product to the buffer.

[0010] In some embodiments, the flow ratio between the first input pipeline and the second input pipeline is used to control the percolation ratio.

[0011] In some embodiments, the flow rate ratio between the first input channel and the second input channel is used to control the buffer exchange ratio.

[0012] In response to the above technical problems, the second aspect of the present invention discloses a control method based on an online concentration system, the steps of which are as follows:

[0013] Step 1: The flow rate of the first input pipeline is F1, and the flow rate of the second input pipeline is F2; ​​the first input pipeline and the second input pipeline respectively deliver buffer solution, and the ratio of F1 to F2 is controlled;

[0014] Step 2: Control the flow rates of the first input pipeline and the second input pipeline so that F1 and F2 reach the target flow rates; at the same time, control the flow rate F3 in the first output pipeline so that F3 is at zero flow or low flow rate;

[0015] Step 3: Turn on the reflux pump on the circulation pipeline and control the flow on the circulation pipeline to achieve the target flow;

[0016] Step 4: When F1 and F2 reach the target flow rate, control the flow rate in the first output pipe so that F3 reaches the target flow rate;

[0017] Step 5: When F3 reaches the target flow rate, the first input pipe is connected to the product tank to deliver the product to be processed; so that the flow rate F1 adjusts itself and stabilizes again;

[0018] Step 6: Continuously and stably control the flow of the first input pipeline, the second input pipeline, the circulation pipeline, and the first output pipeline.

[0019] In response to the above technical problems, the third aspect disclosed in the present invention proposes a chromatography system based on an online concentration system, including the above-mentioned online concentration system, wherein the first output pipe of the online concentration system is connected to the inlet end of the chromatography column through the third input pipe, and the outlet end of the chromatography column is connected to the collection pipe and the waste pipe respectively through the second output pipe, and the inlet ends of the collection pipe and the waste pipe are respectively provided with valves.

[0020] In some embodiments, a valve is provided at the outlet end of the first output pipe of the online replacement system for adjusting the resistance of sample loading.

[0021] In some embodiments, a side branch pipeline is connected between the third input pipeline at the inlet end of the chromatography column and the second output pipeline at the outlet end of the chromatography column. The third input pipeline is provided with a valve, and the valve is located between the inlet end of the chromatography column and the connection part between one end of the side branch pipeline and the second output pipeline; the second output pipeline is provided with two valves, and the two valves are respectively located on both sides of the connection part between the other end of the side branch pipeline and the second output pipeline.

[0022] Beneficial effects of the present invention:

[0023] (1) The present invention provides a circulation pipe between the inlet and intercepting ends of the tangential flow filter. The tangential flow is controlled by a reflux pump. This improves fluidity when the flow rate at the intercepting end outlet of the tangential flow filter is low, thereby preventing protein accumulation on the membrane surface. This is beneficial for increasing yield.

[0024] (2) The flow control of four pumps is significantly improved compared with the existing UF / DF, and the flow control is replaced by pressure control.

[0025] (3) Continuous online buffer exchange and loading of samples during chromatography are performed without relying on traditional TFF batch operations. No circulation tanks, no weight sensors, no stirring devices, and no spray heads are required, thus reducing the equipment footprint and costs.

[0026] (4) With sufficient membrane area, one or half a PFR can achieve the ideal state. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 Schematic diagram of the structure of the concentration system of the present invention.

[0028] Figure 2 Schematic diagram of the structure of the chromatography system of the present invention. DETAILED DESCRIPTION

[0029] The technical content of the present invention is described below using specific embodiments. Those skilled in the art will readily understand the other advantages and effects of the present invention from the disclosure herein. The present invention may also be implemented or applied through other different specific embodiments. Those skilled in the art may make various modifications and alterations without departing from the spirit of the present invention.

[0030] Before introducing the specific embodiments of the present disclosure in detail, some terms used in the present disclosure are first explained.

[0031] Unless otherwise defined hereinafter, the meaning of all technical terms and scientific terms used herein is intended to be the same as those generally understood by those skilled in the art. Mention that the technology used in this article is intended to refer to the technology generally understood in the art, including those changes in technology that are obvious to those skilled in the art or the replacement of equivalent technologies. Although it is believed that the following terms are well understood by those skilled in the art, the following definitions are still set forth to better explain the present invention. When a top-grade product name appears in this article, it is intended to refer to its corresponding top-grade product. All patents, published patent applications and publications cited herein are incorporated herein by reference.

[0032] As used herein, the terms “connect,” “connected,” “coupled,” or similar terms are not limited to direct connections but also include indirect connections.

[0033] As used herein, a "sample" is a biomolecule, including proteins, nucleic acids, lipids, carbohydrates, small nucleotides, amino acids, and their derivatives.

[0034] As used herein, the term "online monitoring" or "real-time monitoring" refers to the real-time detection of certain parameters or properties of a buffer, reaction fluid, or fluid flowing out of a flow reactor, such as pH, pressure, flow rate, conductivity, etc., during the use of a chromatography system. Unlike offline detection or analysis, online monitoring or real-time monitoring can provide real-time feedback of the detection results.

[0035] For example, the positional relationship words "up", "down", "left", "right", "front", "back", etc. involved in this article are determined according to the layout direction of the drawings in the specification. They are only used to express relative positional relationships. When the absolute position of the described object changes, the relative positional relationship may also change accordingly.

[0036] As mentioned in the article, UF / DF, UF (ultrafiltration or concentration) refers to the separation of components based on molecular weight and size to isolate the biological agent of interest. DF (filtration or buffer exchange) refers to the addition of a buffer to replace the liquid surrounding the target with a liquid that is more conducive to storage stability.

[0037] As referred to herein, PFR refers to a tubular reactor.

[0038] Reference Figure 1 As shown, the online concentration system includes a first input pipeline 1 and a second input pipeline 2; a first feed pump 3 is provided on the first input pipeline 1, and a first flow sensor 4 is provided on the first input pipeline 1 at the outlet position of the first feed pump 3, a second feed pump 5 is provided on the second input pipeline 2, and a second flow sensor 6 is provided on the second input pipeline 2 at the outlet position of the second feed pump 5; the output ends of the first input pipeline 1 and the second input pipeline 2 are respectively connected to the inlet end of the mixer 7 through pipelines, the outlet end of the mixer 7 is connected to the inlet end of the tangential flow filter 8 through a pipeline, the intercepting end of the tangential flow filter 8 is connected to the first output pipeline 9, a first discharge pump 10 is provided on the first output pipeline 9, and a third flow sensor 11 is provided on the first output pipeline 9 at the outlet end of the first discharge pump 10; a circulation pipeline 13 is connected between the inlet end and the intercepting end of the tangential flow filter 8, one end of the circulation pipeline 13 is connected to the inlet end of the first discharge pump 10 on the first output pipeline 9, and the other end of the circulation pipeline 13 is connected to the pipeline at the inlet end of the mixer 7. The circulation pipe 13 is provided with a reflux pump 14 , and a fourth flow sensor 15 is provided on the circulation pipe 13 at the outlet end of the reflux pump 14 . The permeate end of the tangential flow filter 8 is connected to the permeate pipe 12 .

[0039] The flow rates of the first feed pump 3 on the first input pipe 1 and the second feed pump 5 on the second input pipe 2 control the transmembrane pressure on the tangential flow filter 8. The reflux pump 14 on the circulation pipe 13 is only used to increase the tangential flow, not the transmembrane pressure.

[0040] The flow rate of the first discharge pump 10 on the first output pipe 9 controls the tangential flow rate of the fluid.

[0041] The first input pipe 1 and the second input pipe 2 are used to transport different fluids; when the first input pipe 1 transports the product to be processed, the second input pipe 2 is used to input the buffer solution; when the first input pipe 1 transports the buffer solution, the second input pipe 2 is used to input the product to be processed.

[0042] During use, the first and second input conduits 1 and 2 are connected to a buffer solution, respectively. The flow rate of the first input conduit 1 under the action of the first feed pump 3 is F1, and the flow rate of the second input conduit 2 under the action of the second feed pump 5 is F2. The ratio of F1 to F2 is controlled. This ratio is used to control the diafiltration ratio; the ratio of F1 to F2 is used to control the buffer exchange ratio.

[0043] The first feed pump 3 and the second feed pump 5 are controlled so that the flow rates of the first input pipeline 1 and the second input pipeline 2 reach the target flow rates; at the same time, the first discharge pump 10 does not work, and the flow rate F3 in the first output pipeline 9 is at zero flow or low flow rate.

[0044] At the same time, the reflux pump 14 on the circulation pipe 13 is turned on. When F1 and F2 reach the target flow rate, the first discharge pump 10 is started to control the flow in the circulation pipe 13 to reach the target flow rate. At the same time, the flow in the first output pipe 9 is controlled so that F3 reaches the target flow rate.

[0045] When the flow rate F3 in the first output pipe 9 reaches the target flow rate, the first input pipe 1 is connected to the product tank to input the product to be processed, so that the flow rate F1 automatically adjusts and stabilizes again. The first feed pump 3, the second feed pump 5, the reflux pump 14, and the first output pump 10 are continuously and stably operated to control the flow rates of the first input pipe 1, the second input pipe 2, the circulation pipe 13, and the first output pipe 9.

[0046] Reference Figure 2 As shown, the chromatography system based on the online concentration system includes a first input pipeline 21 and a second input pipeline 22. The first input pipeline 21 is provided with a first feed pump 23, and a first flow sensor 24 is provided on the first input pipeline 21 at the outlet position of the first feed pump 23. The second input pipeline 22 is provided with a second feed pump 25, and a second flow sensor 26 is provided on the second input pipeline 22 at the outlet position of the second feed pump 25. The output ends of the first input pipeline 21 and the second input pipeline 22 are connected to the inlet end and the outlet end of the mixer 27 respectively. The inlet of the tangential flow filter 28 is connected via a pipeline, and the intercepting end of the tangential flow filter 28 is connected to a first output pipeline 29. A first discharge pump 210 is provided on the first output pipeline 29, and a third flow sensor 211 is provided on the first output pipeline 29 at the outlet of the first discharge pump 210. A circulation pipeline 229 is connected between the inlet and intercepting ends of the tangential flow filter 28. One end of the circulation pipeline 229 is connected to the inlet of the first discharge pump 210 on the first output pipeline 29, and the other end is connected to the pipeline at the inlet of the mixer 27. A reflux pump 230 is provided on the circulation pipeline 229, and a fourth flow sensor 231 is provided on the circulation pipeline 229 at the outlet of the reflux pump 230.

[0047] The permeate end of the tangential flow filter 28 is connected to the permeate line 212. The outlet of the first output line 29 is connected to the inlet of the third input line 214 via a first valve 213. The outlet of the third input line 214 is connected to the inlet of the chromatography column 215, which in turn is connected to the inlet of the second output line 216. The outlet of the second output line 216 is connected to the inlets of the collection line 217 and the waste line 218, respectively. A bypass line 219 is connected between the third input line 214 and the second output line 216. A second valve 220 is provided on the third input line 214 between one end of the bypass line 219 and the connection point between the third input line 214 and the inlet of the chromatography column 215. A third valve 221 and a fourth valve 222 are provided on the second output line 216 on either side of the connection point between the other end of the bypass line 219 and the second output line 216, respectively. A fifth valve 223 is also provided on the bypass line 219.

[0048] The inlet ends of the collecting pipe 217 and the waste discharge pipe 218 are respectively provided with a sixth valve 224 and a seventh valve 225 .

[0049] A UV sensor 226 , a conductivity transmitter 227 and a pH sensor 228 are respectively provided on the second output pipe 216 between one side of the fourth valve 222 and the outlet end of the second output pipe 216 .

[0050] The first valve 213 at the outlet end of the first output pipe 29 is installed based on the size of the sample loading resistance. If the resistance is large, the first valve 213 is not required. If the resistance is small, the first valve 213 needs to be installed to adjust the sample loading resistance.

[0051] During use, the first input conduit 21 and the second input conduit 22 are connected to the buffer solution, respectively. The flow rate of the first input conduit 21 under the action of the first feed pump 23 is F1, and the flow rate of the second input conduit 22 under the action of the second feed pump 25 is F2. The ratio of F1 to F2 is controlled. This ratio is used to control the diafiltration ratio; the ratio of F1 to F2 is used to control the buffer exchange ratio.

[0052] The first feed pump 23 and the second feed pump 25 are controlled so that the flow rates of the first input pipeline 21 and the second input pipeline 22 reach the target flow rates; at the same time, the first discharge pump 210 does not work, and the flow rate F3 in the first output pipeline 29 is at zero flow or low flow.

[0053] At the same time, the reflux pump 230 on the circulation pipe 229 is turned on. When F1 and F2 reach the target flow rate, the first discharge pump 210 is started to control the flow in the circulation pipe 213 to reach the target flow rate. At the same time, the flow in the first output pipe 29 is controlled so that F3 reaches the target flow rate.

[0054] When the flow F3 in the first output pipe 29 reaches the target flow, the buffer solution delivered in the above steps to achieve stable flow delivery is debugged and delivered through the third input pipe 214, the bypass pipe 219, and the second output pipe 216, and then collected by the waste pipe 218.

[0055] At this point, connect the first input pipe 21 to the product tank to feed the product to be processed, causing the flow rate F1 to automatically adjust and stabilize. Continue to operate the first feed pump 23, the second feed pump 25, the reflux pump 230, and the first discharge pump 210 stably to control the flow rates of the first input pipe 21, the second input pipe 22, the circulation pipe 229, and the first discharge pipe 29.

[0056] When the flow rate reaches the chromatographic loading rate, the sample to be processed is transported to the inlet of the chromatography column 215 through the third input pipe 214, and after chromatography, flows out from the outlet of the chromatography column 215. The waste liquid is collected through the waste pipe 218.

Claims

1. Online concentration system, characterized in that, The invention comprises a first input pipeline and a second input pipeline; the first input pipeline and the second input pipeline are respectively provided with a first feed pump and a second feed pump; the output ends of the first input pipeline and the second input pipeline are connected to the inlet end of the tangential flow filter through pipelines; the intercepting end of the tangential flow filter is connected to the first output pipeline; the first output pipeline is provided with a first discharge pump; a circulation pipeline is provided between the inlet end and the intercepting end of the tangential flow filter, and the circulation pipeline is provided with a reflux pump; the permeate end of the tangential flow filter is connected to the permeate pipeline.

2. The online concentration system according to claim 1, characterized in that A mixer is provided between the output ends of the first input pipeline and the second input pipeline and the inlet end of the tangential flow filter, and the outlet of the circulation pipeline is located at the inlet end of the mixer.

3. The online concentration system according to claim 1, characterized in that: In some embodiments, flow sensors are respectively provided on the first input pipe and / or the second input pipe and / or the first output pipe and / or the circulation pipe.

4. The online concentration system according to claim 1, characterized in that The flow rate ratio between the first input pipeline and the second input pipeline is used to control the concentration ratio of the product and the buffer solution.

5. The online concentration system according to claim 1, characterized in that: The flow rate ratio between the first input pipeline and the second input pipeline is used to control the percolation ratio.

6. The online concentration system according to claim 1, characterized in that: The flow rate ratio between the first input channel and the second input channel is used to control the buffer exchange ratio.

7. A control method based on an online concentration system, characterized in that: The steps are: Step 1: The flow rate of the first input pipeline is F1, and the flow rate of the second input pipeline is F2; ​​the first input pipeline and the second input pipeline respectively deliver buffer solution, and the ratio of F1 to F2 is controlled; Step 2: Control the flow rates of the first input pipeline and the second input pipeline so that F1 and F2 reach the target flow rates; at the same time, control the flow rate F3 in the first output pipeline so that F3 is at zero flow or low flow rate; Step 3: Turn on the reflux pump on the circulation pipeline and control the flow on the circulation pipeline to achieve the target flow; Step 4: When F1 and F2 reach the target flow rate, control the flow rate in the first output pipe so that F3 reaches the target flow rate; Step 5: When F3 reaches the target flow rate, the first input pipe is connected to the product tank to deliver the product to be processed; so that the flow rate F1 adjusts itself and stabilizes again; Step 6: Continuously and stably control the flow of the first input pipeline, the second input pipeline, the circulation pipeline, and the first output pipeline.

8. A chromatography system based on an online concentration system, characterized in that The online concentration system comprises the above-mentioned claims 1-6, wherein the first output pipe of the online concentration system is connected to the inlet end of the chromatography column through the third input pipe, the outlet end of the chromatography column is connected to the collection pipe and the waste pipe respectively through the second output pipe, and the inlet ends of the collection pipe and the waste pipe are respectively provided with valves.

9. The chromatography system based on the online concentration system according to claim 8, characterized in that: The outlet end of the first output pipeline of the online replacement system is provided with a valve for adjusting the resistance of sample loading.

10. The chromatography system based on the online concentration system according to claim 8, characterized in that: A side branch pipeline is connected between the third input pipeline at the inlet end of the chromatography column and the second output pipeline at the outlet end of the chromatography column. The third input pipeline is provided with a valve, and the valve is located between the inlet end of the chromatography column and the connection part between one end of the side branch pipeline and the second output pipeline; the second output pipeline is provided with two valves, and the two valves are respectively located on both sides of the connection part between the other end of the side branch pipeline and the second output pipeline.

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