CN113559712A - Tangential flow filtration system and method - Google Patents

Tangential flow filtration system and method Download PDF

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CN113559712A
CN113559712A CN202010355618.5A CN202010355618A CN113559712A CN 113559712 A CN113559712 A CN 113559712A CN 202010355618 A CN202010355618 A CN 202010355618A CN 113559712 A CN113559712 A CN 113559712A
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tangential flow
flow filtration
inlet
pump
stage
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皮健
蒋俊俊
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Wuxi Biologics Shanghai Co Ltd
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Wuxi Biologics Shanghai Co Ltd
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
    • B01D61/14—Ultrafiltration; Microfiltration
    • B01D61/145—Ultrafiltration
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
    • B01D61/14—Ultrafiltration; Microfiltration
    • B01D61/145—Ultrafiltration
    • B01D61/146—Ultrafiltration comprising multiple ultrafiltration steps
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D63/00—Apparatus in general for separation processes using semi-permeable membranes
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D65/00—Accessories or auxiliary operations, in general, for separation processes or apparatus using semi-permeable membranes
    • B01D65/10—Testing of membranes or membrane apparatus; Detecting or repairing leaks

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  • Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Separation Using Semi-Permeable Membranes (AREA)

Abstract

Tangential flow filtration systems and methods are disclosed. The system includes a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux inlet of the tangential flow filtration device.

Description

Tangential flow filtration system and method
Technical Field
The present application relates to tangential flow filtration systems and methods. The system and method of the present application are robust, easy to operate and scale up, and result in stable volume concentrations.
Background
Tangential flow filtration is a membrane separation process that separates the desired product according to molecular weight, molecular size, and other characteristics. In the traditional tangential flow filtration, fermentation liquor circulates to an imported liquid storage tank through a reflux port in the fermentation liquor concentration process, the set volume concentration multiple is achieved by continuously filtering out the fermentation liquor without target protein through a permeation end, and the concentration efficiency is improved by increasing transmembrane pressure (TMP) according to the process requirement. However, the action of shear forces on the pump body and membrane surface over time affects the stability of the target product. For this purpose, Single pass tangential flow filtration (SP-TFF) increases the retention time of the fermentation broth and the membrane by increasing the channel length, achieving a set volume concentration factor at a relatively low transmembrane pressure. Compared with the traditional tangential flow filtration system, the one-way tangential flow filtration system can be suitable for a continuous flow process, the scale of a liquid storage tank in the amplification production is reduced, and the influence of a circulation system on the stability of the protein is reduced.
However, one-way tangential flow filtration still has problems, for example, based on the Millipore SP-TFF device, in the process of concentrating fermentation liquor by one-way tangential flow filtration, the transmembrane pressure rises as the inlet pressure and the reflux port pressure rise, and for pressure control, the reflux port pressure rise will cause the volume concentration multiple to drop along with the pressure rise. In order to keep the transmembrane pressure and the volume concentration multiple stable, xiaoji (research on control of tangential flow and transmembrane pressure of a hollow fiber ultrafiltration membrane system, guangdong chemical engineering, 2011, 8 th, 38 th volume, 220 th total) proposes that a pressure sensor is respectively installed at a backflow port and a permeation end, the transmembrane pressure of the system is obtained through calculation, a pneumatic regulating valve is installed at the backflow port, and a required transmembrane pressure value is obtained by regulating the opening degree of the backflow port regulating valve. However, if the pressure at the return port is controlled and adjusted by the adjusting valve or the flow limiting valve to maintain a stable volume concentration multiple, the whole process needs to be continuously adjusted and monitored, the operation is complex, the volume concentration multiple is unstable, the amplification production is not facilitated, and the optimization of process parameters is also not facilitated. In the process of concentrating the fermentation liquor, along with the continuous increase of the sample loading amount, complex components in the fermentation liquor can form an adsorption layer on the surface of the membrane, so that the flux of the system under unit pressure is reduced, and the stability of the volume concentration multiple is further influenced.
In addition, water flux testing is required before tangential flow filtration. For multi-stage ultrafiltration membranes, the prior art detects the water flux of each stage of membrane separately by changing the flow path (series connection to parallel connection) or splitting each stage of membrane. Such water flux test is cumbersome to operate and is inefficient.
Clearly, there remains a need in the art for a tangential flow filtration system and method that is robust, easy to operate and scale-up, and that results in stable transmembrane pressures and volumetric concentrations. At the same time, there remains a need in the art for a water flux test that is efficient/convenient for large scale tangential flow filtration production.
Disclosure of Invention
To address the above stated problems, one aspect of the present application provides a tangential flow filtration system comprising a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux outlet of the tangential flow filtration device.
In one embodiment of the present application, the backflow control device includes a backflow pump and/or a backflow end valve. In one embodiment of the application, the return flow control device comprises a return pump and a return end valve, the return pump and the return end valve being in parallel or switchable with each other. In one embodiment of the present application, the backflow control device includes a backflow pump or a backflow end valve, and the backflow pump and the backflow end valve are replaceable. In one embodiment of the present application, the return pump and the return end valve are switched to each other by adjusting the communication state of the pipeline. In one embodiment of the present application, the reflux pump is in parallel with the reflux end valve and the liquid flow is controlled by opening, adjusting or closing the function of the reflux pump and/or the reflux end valve. In one embodiment of the present application, the back-flow pump and back-flow end valve are physically replaceable, including, for example, being removed and installed. In one embodiment of the present application, the tangential flow filtration system further comprises a reservoir in fluid communication with the inlet pump and a collection device in fluid communication with the return flow control device. In one embodiment of the present application, the fluid reservoir device is a fluid reservoir bottle or a fluid reservoir tank. In one embodiment of the present application, the collection device is a collection bottle or a collection canister. In one embodiment of the present application, the reservoir contains a liquid selected from the group consisting of: sample and buffer. In one embodiment of the present application, the sample is a perfusion culture sample. In one embodiment of the present application, the sample is a protein solution. In one embodiment of the present application, the protein is an antibody. In one embodiment of the present application, the antibody is a monoclonal antibody. In one embodiment of the present application, the sample is a fermentation broth supernatant. In another embodiment of the present application, the fermentation broth is cleaned from an animal cell fermentation broth supernatant, a plant cell fermentation broth supernatant, or a microbial cell fermentation broth supernatant. In another embodiment of the present application, the fermentation broth supernatant is a fermentation broth supernatant of chinese hamster ovary cells or mouse myeloma cells. In another embodiment of the present application, the fermentation broth supernatant is a fermentation broth supernatant of chinese hamster ovary cells or mouse myeloma cells into which the foreign gene is integrated. In another embodiment of the present application, the cleaning on the fermentation broth is from: cell culture supernatant after batch fed-batch culture and clarification process treatment, or cell culture filtrate after perfusion culture. In one embodiment of the present application, the buffer is selected from the group consisting of an equilibration buffer, a regeneration buffer, a rinse buffer, a disinfection buffer, or a preservation solution.
In one embodiment of the present application, the tangential flow filtration device is a single pass tangential flow filtration device. In one embodiment of the present application, the tangential flow filtration device comprises one or more ultrafiltration membrane packages. In one embodiment of the present application, the tangential flow filtration device comprises a series of multi-stage ultrafiltration membrane packages. In one embodiment of the present application, the tangential flow filtration device comprises three ultrafiltration membrane packages in series. In one embodiment of the present application, the membrane ratio of the tertiary ultrafiltration membrane package is 1: 1. In one embodiment of the present application, the inlet pump and the return pump are independently selected from diaphragm pumps or peristaltic pumps. In another embodiment of the present application, the inlet pump is a diaphragm pump and the return pump is a peristaltic pump. In one embodiment of the present application, the return end valve is a regulator valve. In one embodiment of the present application, the return end valve is a pressure valve.
Another aspect of the present application provides a method of determining tangential flow filtration parameters, comprising: a) determining the transmembrane pressure upper limit of a first-stage membrane; b) constructing a tangential flow filtration system comprising a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux inlet of the tangential flow filtration device; c) determining an upper inlet flux limit; and d) determining the upper limit of the volume concentration factor.
In one embodiment of the present application, determining the first stage transmembrane pressure upper limit is based on a system steady state. In one embodiment of the present application, the system steady state is a condition that avoids severe polarization at the membrane surface, resulting in a change in permeate flux. In one embodiment of the present application, determining the upper first stage transmembrane pressure limit comprises determining a maximum first stage transmembrane pressure attainable before polarization of the first stage membrane occurs. In one embodiment of the present application, the first stage membrane transmembrane pressure ceiling varies with the tangential flow filtration device and the difference in the samples processed. In one embodiment of the present application, the first stage membrane transmembrane pressure upper limit is 12 psi. In one embodiment of the present application, determining the first stage membrane transmembrane pressure ceiling further comprises constructing a tangential flow filtration system comprising a tangential flow filtration device comprising a first stage membrane. In one embodiment of the present application, the first stage membrane in step a) is the same as the first stage membrane of the tangential flow filtration device in step b). In one embodiment of the present application, the tangential flow filtration system further comprises an inlet pump in fluid communication with the inlet of the tangential flow filtration device, and a reflux control device in fluid communication with the reflux inlet of the tangential flow filtration device. In one embodiment of the present application, the return flow control device includes a return end valve.
In one embodiment of the present application, determining the inlet flux upper limit is based on the actual system capacity and the first stage membrane transmembrane pressure upper limit. In one embodiment of the present application, determining the upper inlet flux limit is based on maintaining a maximum inlet flux determined for a steady state of the system. In one embodiment of the present application, determining the upper inlet flux limit comprises determining a maximum inlet flux that achieves an actual system capacity at a fixed volume concentration factor without exceeding the upper first stage membrane transmembrane pressure limit.
In one embodiment of the present application, determining the upper volume concentration factor is based on the actual system loading and the upper first stage membrane transmembrane pressure limit. In one embodiment of the present application, determining the upper volume concentration factor is based on maintaining a maximum volume concentration factor determined at steady state in the system. In one embodiment of the present application, determining the upper limit on volume concentration factor comprises determining a maximum volume concentration factor that achieves an actual system loading at a fixed inlet flux without exceeding the upper limit on first stage membrane transmembrane pressure. In one embodiment of the present application, further comprising determining a lower volume concentration factor. In one embodiment of the present application, determining the lower volume concentration factor is based on large scale production requirements. In one embodiment of the present application, determining the lower limit of the volume concentration factor is based on the concentration factor required for large scale production.
In one embodiment of the present application, determining a system capacity cap is further included. In one embodiment of the present application, determining the upper system loading is based on the upper first stage membrane transmembrane pressure.
In one embodiment of the present application, further comprising performing a system water flux test. In one embodiment of the present application, the system water flux test precedes step a) and/or step c). In one embodiment of the present application, the system water flux test comprises determining the system water flux of the tangential flow filtration device and comparing it to a reference system water flux. In one embodiment of the present application, the reference system water flux is the initial system water flux of the tangential flow filtration device or the system water flux measured in a previous use. In one embodiment of the present application, where the reference system water flux is the initial system water flux, the system water flux is reduced by no more than 20% as compared to the reference system water flux. In one embodiment of the present application, where the reference system water flux is the system water flux determined in a previous use, the system water flux is reduced by no more than 10% compared to the reference system water flux.
Yet another aspect of the present application provides a method of performing tangential flow filtration, comprising: a) constructing a tangential flow filtration system comprising a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux inlet of the tangential flow filtration device; b) setting parameters; and c) operating the system.
In one embodiment of the present application, further comprising performing a system water flux test. In one embodiment of the present application, the system water flux test precedes step b) or step c).
In one embodiment of the present application, the parameters include actual system loading, first stage membrane transmembrane pressure upper limit, inlet flux and volume concentration factor.
Yet another aspect of the present application provides a system water flux testing method, comprising: a) constructing a tangential flow filtration system comprising a tangential flow filtration device comprising a multi-stage ultrafiltration membrane module; b) adding water and operating the tangential flow filtration system; c) adjusting the inlet pressure and the outlet pressure; and d) determining the water flux of the system.
In one embodiment of the present application, the multi-stage ultrafiltration membrane module is a series of multi-stage ultrafiltration membrane modules. In one embodiment of the present application, the multi-stage ultrafiltration membrane module is a series of three stage ultrafiltration membrane modules. In one embodiment of the present application, the membrane ratio of the tertiary ultrafiltration membrane package is 1: 1.
In one embodiment of the present application, the tangential flow filtration system further comprises an inlet pump in fluid communication with the inlet of the tangential flow filtration device, and a return port valve in fluid communication with the return port of the tangential flow filtration device.
In another embodiment of the present application, the tangential flow filtration system comprises a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux outlet of the tangential flow filtration device. In one embodiment of the present application, the reflux control device includes a reflux port valve and optionally a reflux pump, wherein the inlet pressure is regulated by the inlet pump and the outlet pressure is regulated by the reflux port valve when conducting a system water flux test.
In one embodiment of the present application, the inlet pressure and the outlet pressure are adjusted to determine the system transmembrane pressure. In one embodiment of the present application, the system transmembrane pressure is not less than 0.5 bar. In one embodiment of the present application, the system transmembrane pressure is 0.5-1.0 bar. In one embodiment of the present application, the system transmembrane pressure is about 0.8 bar.
Compared with the prior art, the tangential flow filtration system and the method are easy to operate and amplify, parameters of the tangential flow filtration system can be accurately controlled, and stable transmembrane pressure and volume concentration multiple can be realized. Moreover, the system water flux testing method can efficiently/conveniently obtain data results.
Drawings
The present application is described in more detail below with reference to the attached drawing figures, wherein:
FIG. 1a is a schematic view of one embodiment of the tangential flow filtration system of the present application. Wherein, showing the liquid storage bottle, the inlet pump, the one-way tangential flow filter device, the reflux pump, the reflux end valve, the collecting bottle and the pipeline connecting the above elements, the pipeline comprises a pressure gauge.
Fig. lb is a schematic view of an embodiment of a single pass tangential flow filtration device of the present application. Wherein, show first level membrane, second grade membrane, tertiary membrane and connect the pipeline of above component, including the manometer in the pipeline.
Figure 2a is the trend of TMP and volume concentration with the amount of sample in the reflux valve control mode.
FIG. 2b is a pressure trend of the SP-TFF system in a return port valve control mode.
FIG. 3a is a graph showing the trend of TMP and volume concentration with loading for the dual pump control system.
FIG. 3b is the pressure variation trend of the SP-TFF system under the dual-pump control system.
Figure 4 is a graph of water flux as a function of system TMP.
Figure 5 is a graph of simulated first stage membrane Qp versus TMP variation at different inlet fluxes.
Figure 6 is a graph of simulated tertiary membrane Qp versus TMP variation at different inlet fluxes.
Fig. 7a is a graph of pressure versus time.
Figure 7b is a graph of the change in permeate end and outlet flow rates.
Figure 7c is the volume concentration times and the pressure change of each membrane and system across the membrane.
Figure 8 is the trend of 4-fold volumetric concentration system TMP with loading.
Figure 9 is the trend of the 6-fold volume concentration system TMP with loading.
Figure 10 is the trend of 8-fold volumetric concentration system TMP with loading.
Figure 11 is the trend of system TMP with load at 36LMH inlet flux.
Figure 12 is the trend of system TMP with load at 33LMH inlet flux.
Figure 13 is the trend of system TMP with load at 30LMH inlet flux.
Detailed Description
Definition of
In this application, "single pass tangential flow filtration (SP-TFF)" refers to a tangential flow filtration technique in which liquid is passed through a tangential flow filtration device once, rather than being circulated through the device. "Single pass tangential flow filtration device" means a device for single pass tangential flow filtration, the selection of the type being within the routine skill of one skilled in the art.
In this application, the "system transmembrane pressure" is equal to the sum of the inlet and outlet pressures divided by 2 minus the sum of the permeate end pressures of the stages.
In this application, the "first stage membrane transmembrane pressure" is equal to the sum of the inlet pressure and the return port 1 pressure divided by 2 minus the permeate port 1 pressure.
In this application, the "second stage transmembrane pressure" is equal to the sum of the return port 1 pressure and the return port 2 pressure divided by 2 minus the permeate port 2 pressure.
In this application, the "third stage transmembrane pressure" is equal to the sum of the return port 2 pressure and the outlet pressure divided by 2 minus the permeate port 3 pressure.
In the present application, "membrane area" refers to the sum of the total areas of the membranes of the various stages in the membrane package.
In this application, "membrane proportioning" refers to the ratio of the areas of the membranes of each stage in a multi-stage membrane system.
In this application, "inlet flux" is equal to the flow rate of liquid through the inlet divided by the membrane area, which is given in LMH (L/h m)2)。
In this application, the return port of the last stage membrane is also referred to as the "outlet". "Outlet flux" is equal to the total volume of liquid passing through the outlet divided by the membrane area divided by the passage time, in LMH (L @)h*m2)。
In this application, the "permeate end flux (Qp)" is equal to the flow rate of liquid through the permeate end divided by the membrane area of the corresponding membrane of the permeate end, in units of LMH (L/h m)2)
In this application, "system water flux (NWP)" means the volume of water passing through each membrane stage per unit of membrane area per unit of time and per unit of pressure, and is expressed in L/h m2*psi。
In this application, "Volume Concentration Factor (VCF)" refers to the factor by which the volume is reduced in a concentrating operation, and is equal to the inlet flux divided by the outlet flux.
In the present application, the term "system steady state" means that the pressure and corresponding flow velocity distribution of each membrane stage of the system are in a steady state, i.e., the pressure and flow velocity of each membrane stage are not significantly increased or decreased.
As used herein, "system loading" refers to the capacity of a tangential flow filtration system to process, expressed as volume of liquid processed per unit membrane area (L/m)2)。
In this application, the "actual system load" is equal to the volume of the sample to be filtered divided by the membrane area.
In one embodiment of operation, as shown in FIG. 1a, the tangential flow filtration system employs a single pass tangential flow filtration device. During operation, liquid from the liquid storage bottle enters the inlet pump through a pipeline, the liquid enters the one-way tangential flow filtering device through the inlet of the pipeline through the pumping of the inlet pump, permeate liquid leaves the one-way tangential flow filtering device through the permeation end, and trapped liquid leaves the one-way tangential flow filtering device through the backflow port, then when the pumping function of the backflow pump is opened and the backflow end valve is closed so that the liquid cannot pass through the backflow end valve, the trapped liquid enters the backflow pump through the pipeline, finally the trapped liquid enters the collecting bottle through the pipeline through the backflow pump through the pumping of the backflow pump, when the pressure regulating function of the backflow end valve is opened and the backflow pump is closed so that the liquid cannot pass through the backflow pump, the trapped liquid enters the backflow end valve through the pipeline, and finally the trapped liquid enters the collecting bottle through the backflow end valve through the pipeline.
In one embodiment of operation, as shown in fig. 1b, the tangential flow filtration system employs a single pass tangential flow filtration device comprising three ultrafiltration membrane modules in series, in operation, liquid enters the inlet of the first membrane module through a conduit, permeate exits the first membrane module through permeate port 1 and retentate exits the first membrane module through return port 1 and enters the inlet of the second membrane module through a conduit, permeate exits the second membrane module through permeate port 2 and retentate exits the second membrane module through return port 2 and enters the inlet of the third membrane module through a conduit, permeate exits the third membrane module through permeate port 3 and retentate exits the third membrane module through the outlet.
The technical solutions of the present application will be described in detail and fully with reference to the accompanying drawings, and it should be understood that the described embodiments are only some embodiments of the present application, but not all embodiments. All other embodiments obtained by a person of ordinary skill in the art based on the embodiments in the present application without any inventive step are within the scope of protection of the present application.
Example 1
System water flux (NWP)
In order to obtain the water flux data of the three-stage membrane series system more quickly, a method for integrally testing the water flux of the three-stage membrane series system is adopted, namely, an inlet pump and a return end valve are controlled to reach different system transmembrane pressures, and the robustness of the system water flux under different system transmembrane pressures is monitored.
A single-pass tangential flow filtration system is constructed, wherein a Millipore SP-TFF system (Pellicon 3Cassette P3C030C00, Ultracel 30kDa, C flow channel, membrane ratio of 1: 1) is adopted as a single-pass tangential flow filtration device, an inlet pump (diaphragm pump, QF150S (Quattro)) is arranged at an inlet of the device, a return end valve is arranged at an outlet of the device, a liquid storage bottle, the inlet pump, the single-pass tangential flow filtration device, the return end valve and a collecting bottle are sequentially connected by pipelines, and pressure gauges are arranged at the inlet and a return port and a permeation end of each stage of membrane.
And pumping deionized water by an inlet pump, adjusting the inlet pressure to be 0.6bar by the inlet pump, adjusting the outlet pressure to be 0bar by a backflow end valve, enabling the transmembrane pressure of the system to be 0.3bar, running for 5 minutes, and recording the permeation end flow rate, the outlet flow rate, the inlet pressure, the outlet pressure, the permeation end pressure and the water temperature after the membrane pressures and the water temperatures are stable. NWP was calculated using the following formula:
Figure BDA0002473329230000081
wherein P is the permeate end flow rate; k is a temperature correction coefficient; pin ═ inlet pressure; pout ═ outlet pressure; pp — trans end pressure; and a ═ membrane area.
NWP temperature correction coefficient K
T(°F) T(℃) K T(°F） T(℃) K T(°F) T(℃) K
125.6 52 0.595 96.8 36 0.793 68.0 20 1.125
123.8 51 0.605 95.0 35 0.808 66.2 19 1.152
122.0 50 0.615 93.2 34 0.852 64.4 18 1.181
120.2 49 0.625 91.4 33 0.842 62.2 17 1.212
118.4 48 0.636 89.6 32 0.859 60.8 16 1.243
116.6 47 0.647 87.8 31 0.877 59.0 15 1.276
114.8 46 0.658 86.0 30 0.896 57.2 14 1.310
113.0 45 0.670 84.2 29 0.915 55.4 13 1.346
111.2 44 0.682 82.4 28 0.935 53.6 12 1.383
109.4 43 0.694 80.6 27 0.956 51.8 11 1.422
107.6 42 0.707 78.8 26 0.978 50.0 10 1.463
105.8 41 0.720 77.0 25 1.000 48.2 9 1.506
104.0 40 0.734 75.2 24 1.023 46.4 8 1.551
102.2 39 0.748 73.4 23 1.047 44.6 7 1.598
100.4 38 0.762 71.6 22 1.072 42.8 6 1.648
98.6 37 0.777 69.8 21 1.098 41.0 5 1.699
Then, the outlet pressure is adjusted to 2, 3, 4, 5 and 6psi through the return end valve in sequence, and then the inlet pressure is adjusted through the inlet pump, so that the transmembrane pressure of the system is 0.3 bar. If the outlet flow rate drops to 0, the outlet pressure does not increase any more. Each measurement was repeated twice.
The process was then repeated except that the transmembrane pressure of the system was controlled to be 0.5bar, 0.8bar and 1.0bar in that order.
The results are shown in fig. 4, and the higher the transmembrane pressure of the system is, the more stable the water flux test result of the system is. Meanwhile, if the outlet pressure is not controlled to be 0, the deviation of the water flux data relative to other data is large and is close to 10%. In order for the method to test the robustness of the water flux of the system, the outlet pressure needs to be controlled to be greater than 0. The transmembrane pressure of the system used in the water flux test of the system was in the range 0.5-1.0bar, taking into account the acceptable range of 10% deviation for the multiple measurements.
At present, the traditional water flux testing method for the ultrafiltration membrane package is to test the water flux of a single-stage membrane, but to test the water flux of a multi-stage system, a system in a three-stage membrane series mode needs to be changed to be in a three-stage membrane parallel mode, and then the water flux is detected by using a conventional testing mode, or the three-stage series system is directly disassembled to detect the water flux in a grading mode. The invention firstly adopts the water flux test to the whole three-stage membrane series system without changing the connection mode of the three-stage membrane or respectively carrying out the water flux test to the single-stage membrane, thereby completing the water flux test of the system at one time, simplifying the water flux test process and obtaining the water flux data more quickly.
Example 2
Comparison of effects of double-pump control strategy and backflow end valve control strategy
A single-pass tangential flow filtration system is constructed, wherein a Millipore SP-TFF system (Pellicon 3Cassette P3C030C00, Ultracel 30kDa, C flow channel, membrane ratio of 1: 1) is adopted as a single-pass tangential flow filtration device, an inlet pump (a diaphragm pump, QF150S (Quattro)) is arranged at the inlet of the device, a reflux pump (a peristaltic pump, BT100-2J (LongerPump)) and a reflux end valve are arranged at the outlet of the device in parallel, a liquid storage bottle, the inlet pump, the single-pass tangential flow filtration device, the reflux pump, the reflux end valve and a collecting bottle are sequentially connected by pipelines, and pressure gauges are arranged at the inlet and the reflux end and the permeation end of each stage of membrane.
(1) Return end valve control strategy
The system was sterilized by flushing 20L/M with 0.1M NaOH at an inlet flux of 100LMH2After that, the system was cycled for 60 minutes.
The system was tested for systemic water flux as described in example 1 (using a system transmembrane pressure of 0.8 bar), equilibrated at an inlet flux of 100LMH, and 20L/m using an equilibration buffer (50mM Tris-HAc, 150mM NaCl, pH 7.4)2The volume of the system is washed, the pH and the conductance of the effluent are detected until the pH and the conductance of the effluent are equal to a balance buffer solution so as to confirm that the balance of the three-stage ultrafiltration membrane package is completed, a reflux end valve is opened to regulate the pressure and the reflux pump is closed to ensure that liquid cannot pass through the reflux pump, a sample (fermentation broth supernatant of mouse myeloma cells, which is obtained by perfusion culture and clarification process treatment) is pumped into the system, the system is operated, the pump speed of an inlet pump is regulated to a preset inlet flux (36LMH), after 5 minutes of operation, the reflux end valve is regulated to a preset outlet pressure (3psi, the system can reach 4 times of volume concentration multiple by keeping the outlet pressure of 3psi at the moment according to the development data of the previous stage), and the inlet flow rate, the outlet flow rate, the inlet pressure, the reflux port 1 pressure, the reflux port 2 pressure, the outlet pressure, the permeation port 1 pressure, the permeation port 2 pressure and the permeation port 3 pressure are recorded along with time, the transmembrane pressure and the volume concentration factor of the system and each membrane stage are calculated, and the results are shown in fig. 2a and 2b respectively.
(2) Dual pump control strategy
After the sterilization, NWP determination and balance of the three-stage ultrafiltration membrane module are completed as described in (1), the pump function of the reflux pump is opened and the reflux end valve is closed to prevent the liquid from passing through the reflux end valve, the same product is pumped into the system to operate the system, the inlet pump speed is adjusted to a preset inlet flux (36LMH), after 5 minutes of operation, the volume concentration multiple is reached (4 times) by adjusting the reflux pump to a preset outlet flux (9LMH), and the inlet flow rate, the outlet flow rate, the inlet pressure, the reflux port 1 pressure, the reflux port 2 pressure, the outlet pressure, the permeation end 1 pressure, the permeation end 2 pressure and the permeation end 3 pressure are recorded over time, and the transmembrane pressure and the volume concentration multiple of the system and each stage of membranes are calculated, and the results are respectively shown in fig. 3a and 3 b.
In the return end valve control strategy, control of the outlet pressure, which is adjusted based on earlier developed data, can result in drastic changes in the volume concentration factor. The outlet pressure needs to be controlled to be about a certain value for controlling a certain concentration multiple, but actually, the outlet pressure is set to be higher than the preset concentration multiple, which is also an uncertain factor caused by the control of the return end valve, namely, developed data is different from actual operation. Therefore, the control strategy of the return end valve needs to be adjusted correspondingly according to the actual volume concentration multiple, and the previous development data can not provide actual production reference. In addition, as the pressure increases, further attempts to reduce the system pressure will result in dramatic changes in the volume concentration factor. Control of the return end valve complicates production control and requires adjustment of the outlet pressure to the change in the concentration factor at all times. Compared with the control method, the control strategy is simplified by the double-pump control strategy, the pressure of the backflow port does not need to be adjusted constantly according to the change of the concentration multiple, and the long-term stable control on the volume concentration multiple and the pressure can be realized by the arrangement of the backflow pump.
Example 3
The sample is 200L fermentation liquor (fermentation liquor supernatant of Chinese hamster ovary cells, wherein the Chinese hamster ovary cells are integrated with exogenous genes to express monoclonal antibodies, the monoclonal antibodies are obtained by fed-batch culture and clarification process, and the concentration of the monoclonal antibodies in the fermentation liquor supernatant is 0.2mg/mL), in order to realize downstream continuous production, the optimized ranges of import flux and volume concentration multiple are obtained to reduce the volume of the fermentation liquor, reduce the scale of 200L downstream production, reduce the usage amount of downstream affinity fillers and reduce the cost.
The first stage is as follows: determining the upper limit of the transmembrane pressure of the first-stage membrane
In a three-stage ultrafiltration membrane package system, because the volume of the feed liquid processed by a first-stage membrane is the largest, and the concentration of the feed liquid processed by a third-stage membrane is the highest, the two extreme conditions are taken as research objects in the simulation experiment, and the purpose is to find the upper limit of the transmembrane pressure of the first-stage membrane and the upper limit of the transmembrane pressure of the third-stage membrane, which avoid the serious polarization phenomenon on the surface of the membrane package so as to enable the system to be in a stable state.
The same single pass tangential flow filtration system as in example 2 was used except a single stage ultrafiltration membrane pack (P3C030C00, Ultracel 30kDa, C flow channel) was used.
TMP was optimized for simulated first stage membranes: TMP-Qp curves were plotted. Samples were used to simulate the first stage membrane treatment feed. After the system was disinfected, NWP measured and balanced as described in example 2, the reflux port valve was opened to regulate pressure and the reflux pump was closed to stop the passage of liquid through the reflux pump, the sample was pumped into the system to circulate the system under the current sample, the inlet pump speed was adjusted to the preset inlet flux (240 LMH respectively), after 5 minutes of operation, the reflux port pressure was adjusted by adjusting the reflux port valve to bring the transmembrane pressure from the preset lower pressure to the maximum pressure (3, 5, 10, 15, 20 and 25psi in order from lower to maximum), and the permeation port flux was calculated by recording the permeation port flow rates at different transmembrane pressures. The inlet flux was then adjusted to the next preset value (180, 120 and 60LMH) and the above steps were repeated. The results for transmembrane pressure and permeate end flux at different inlet fluxes are shown in figure 5.
TMP was optimized for simulated tertiary membrane: TMP-Qp curves were plotted. The sample concentrated to 3 times the concentration was used to simulate the feed solution for the third stage membrane treatment, using the same procedure as for simulating the first stage membrane to optimize TMP, and the results are shown in fig. 6.
In a single-pass tangential flow filtration system, the inlet flux to the first and third stage membranes and the concentration factor of the feed being treated are different, which reduces the flux of the third stage membrane. In the system, the three-stage membrane TMP is smaller than the first stage and the second stage, but the processed feed liquid is processed by the first stage membrane and the second stage membrane, so that the upper limit of the transmembrane pressure of the first stage membrane is firstly reached according to the change of the system pressure, and the transmembrane pressure of the first stage membrane is used as a judgment standard for judging the polarization of the membrane surface and the adsorption of protein.
For the first stage membrane, at each preset inlet flux, the permeate side flux initially increased with increasing TMP, but after reaching a certain value, the permeate side flux no longer increased due to the polarization of the membrane surface and the increase in protein adsorption even though TMP continued to rise. Similar trends were also observed for the third stage membrane. Therefore, it is desirable to control the first stage membrane TMP upper limit, i.e., the system loading endpoint. As shown in fig. 5 and 6, the upper transmembrane pressure limit of the first stage membrane was set at 12psi in order to reduce polarization of the membrane surface and adsorption of proteins.
And a second stage: determining an upper inlet flux limit
To reduce the rate of adsorption of impurities on the membrane package surface, maintaining a relatively long throughput, we optimized the inlet flux of the membrane package. The treatment speed of the membrane package is improved by improving the inlet flux, and the condition of adsorbing impurities on the surface of the membrane is judged according to the time length of the system maintaining the steady state.
Adopting a one-way tangential flow filtration system which is the same as that in the embodiment 2, after the disinfection, NWP measurement and balance of a three-stage ultrafiltration membrane package are completed, opening the pumping function of a reflux pump and closing a reflux end valve to prevent liquid from passing through the reflux end valve, pumping a sample into the system, adjusting the pumping speed of an inlet pump to a preset inlet pressure (5 psi, 8 psi, 10psi, 12psi, 15 psi, 18 psi and 20psi from low to high), continuously operating for the same time interval (1800s), recording the inlet pressure, the pressure of a reflux port 1, the pressure of a reflux port 2, the outlet pressure, the inlet flow rate, the flow rate of a permeation end 1, the flow rate of a permeation end 2, the flow rate of a permeation end 3 and the outlet flow rate along with time, adjusting the pumping speed of the inlet pump to other preset pressure values, continuously operating for the same time interval respectively and recording the parameters in turn, calculating the inlet flux, the outlet flux and the volume concentration multiple, the results are shown in FIGS. 7 a-c.
The steady state of the system is judged based on the change of the pressure and the flow rate of each port, and when the inlet flux is increased to enable the inlet pressure to be at 10psi, the corresponding inlet flux is 36LMH, and the system can maintain the pressure and the flow rate of each port to be steady, namely, the steady state of the system in a relatively long time. When the inlet pressure reaches 12psi or higher, the individual port pressures and flow rates no longer remain stable over time, and the volume concentration factor no longer stabilizes, at which point the system is in an unstable state. This indicates that after the inlet pressure reached 12psi, the system was no longer in a steady state condition, which would benefit the ability of the system to handle feed. Thus, to bring the system to system steady state, the inlet pressure is set to no greater than 10psi, corresponding to an inlet flux of no greater than 36 LMH.
And a third stage:
(1) volume concentration factor versus system loading
Based on the results of the second stage, we first selected a dual pump system to maintain a relatively long inlet pressure of 10psi, and the system termination conditions are given according to the first stage experiments, in which case the system will terminate the system process at a first stage membrane transmembrane pressure of 12 psi. The relationship of the volume concentration factor to the system loading was investigated under this condition.
Adopting a one-way tangential flow filtration system which is the same as that in the embodiment 2, after the disinfection, NWP measurement and balance of a three-stage ultrafiltration membrane package are completed, opening the pumping function of a reflux pump, closing a reflux end valve to prevent liquid from passing through the reflux end valve, pumping a sample into the system, adjusting the pumping speed of an inlet pump to keep the inlet flux at 36LMH, after 5 minutes of operation, adjusting the pumping speed of the reflux pump to keep the outlet flux at 9LMH (namely 4 times VCF), continuously operating until the sample is exhausted or the first-stage transmembrane pressure reaches 12psi, recording the inlet pressure, the reflux port 1 pressure, the reflux port 2 pressure, the outlet pressure, the permeation end 1 flow rate, the permeation end 2 flow rate, the permeation end 3 flow rate and the outlet flow rate at the same time interval, and calculating the system loading capacity, the inlet flux, the outlet flux, the volume concentration multiple and the transmembrane pressure of each stage of membrane. The above experiment was repeated, each time adjusting the pump speed of the reflux pump to maintain the outlet flux at 6LMH and 4.5LMH, respectively (to achieve 6 and 8 times VCF, respectively). The volume concentration fold versus transmembrane pressure change as a function of system loading was plotted as shown in figures 8, 9 and 10 based on the recorded data.
As shown in FIGS. 8, 9 and 10, at an inlet flux of 36LMH, 4, 6 and 8 VCFs can achieve about 150L/m, respectively2About 75L/m2And about 40L/m2The system capacity of (c). The larger the volume concentration factor, the lower the system loading.
(2) Relationship between import flux and system capacity
Based on the results of (1), we first selected a dual pump system that maintained 6 VCF for a relatively long time, and the system termination conditions are given according to the first stage experiment, in which case the system would terminate the system process at a first stage membrane transmembrane pressure of 12 psi. The relationship of the import flux to the system capacity was studied under this condition.
Adopting a one-way tangential flow filtration system which is the same as that in the embodiment 2, after the disinfection, NWP measurement and balance of a three-stage ultrafiltration membrane package are completed, opening the pumping function of a reflux pump, closing a reflux end valve to prevent liquid from passing through the reflux end valve, pumping a sample into the system, adjusting the pumping speed of an inlet pump to keep the inlet flux at 36LMH, after 5 minutes of operation, adjusting the pumping speed of the reflux pump to keep the outlet flux at 6LMH (namely 6 times VCF), continuously operating until the sample is exhausted or the first-stage transmembrane pressure reaches 12psi, recording the inlet pressure, the reflux port 1 pressure, the reflux port 2 pressure, the outlet pressure, the permeation end 1 flow rate, the permeation end 2 flow rate, the permeation end 3 flow rate and the outlet flow rate at the same time interval, and calculating the system loading capacity, the inlet flux, the outlet flux, the volume concentration multiple and the transmembrane pressure of each stage of membrane. The above experiment was repeated, each time the pump speeds of the inlet pump and the reflux pump were simultaneously adjusted so that the inlet flux and the outlet flux were maintained at 33LMH and 5.5LMH (fig. 12) and 30LMH and 5LMH (fig. 13), respectively. The volume concentration fold versus transmembrane pressure change as a function of system loading was plotted as shown in figures 11, 12 and 13 based on the recorded data.
As shown in FIGS. 11, 12 and 13, at 6 times VCF, 36LMH, 33LMH and30LMH can respectively realize about 75L/m2About 90L/m2And more than 100L/m2The system capacity of (c). The greater the inlet flux, the lower the system loading.
Based on the above experiments, the ultimate goal was to obtain a range of inlet flux and volume concentration factors that achieve practical system loading in a tangential flow filtration system.
The above are only examples of the present application, and do not limit the scope of the present application. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. All embodiments need not be described or illustrated herein. The technical solutions like this formed by equivalent transformation or equivalent substitution fall within the protection scope of the present application.

Claims (19)

1. A tangential flow filtration system comprising a tangential flow filtration device, an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux outlet of the tangential flow filtration device.
2. The system of claim 1, the backflow control device comprising a backflow pump and/or a backflow end valve.
3. The system of claim 2, the backflow control device comprising a backflow pump and a backflow end valve, the backflow pump and the backflow end valve being in parallel or switchable with each other.
4. The system of claim 2, the backflow control device comprising a backflow pump or a backflow end valve, the backflow pump and the backflow end valve being interchangeable.
5. The system of claim 1, wherein the tangential flow filtration device is a single pass tangential flow filtration device, and/or the inlet pump and the reflux pump are independently selected from a diaphragm pump or a peristaltic pump.
6. The system of claim 1, wherein the tangential flow filtration device comprises one or more stages of ultrafiltration membrane packages, preferably a series of multi-stage ultrafiltration membrane packages, more preferably a series of three stage ultrafiltration membrane packages.
7. The system of claim 1, further comprising a reservoir in fluid communication with the inlet pump and a collection device in fluid communication with the return flow control device.
8. A system according to claim 7, the reservoir containing a liquid selected from the group consisting of: a sample, preferably a protein solution or supernatant of a cell broth, more preferably a supernatant of an animal cell broth, most preferably a supernatant of a broth of chinese hamster ovary cells or mouse myeloma cells, or a buffer.
9. A method of determining tangential flow filtration parameters, comprising:
a) determining the transmembrane pressure upper limit of a first-stage membrane;
b) constructing a tangential flow filtration system of any one of claims 1-8;
c) determining an upper inlet flux limit; and
d) the upper limit of the volume concentration factor is determined.
10. The method of claim 9, wherein the determination of step a) is based on system steady state.
11. The method of claim 9, the determining of step a) further comprising constructing a tangential flow filtration system comprising a tangential flow filtration device comprising a first stage membrane that is the same as the first stage membrane of the tangential flow filtration device of step b).
12. The method of claim 9, wherein the determination of step c) is based on the actual system loading and the upper first stage membrane transmembrane pressure limit determined in a).
13. The method of claim 9, wherein the determination of step d) is based on the actual system loading and the upper first stage membrane transmembrane pressure limit determined in a).
14. A method of performing tangential flow filtration comprising:
a) constructing a tangential flow filtration system of any one of claims 1-8;
b) setting parameters; and
c) the system is operated.
15. The method of claim 14, the parameters comprising actual system loading, first stage membrane transmembrane pressure upper limit, inlet flux, and volume concentration factor.
16. A system water flux testing method comprising:
a) constructing a tangential flow filtration system comprising a tangential flow filtration device comprising a multi-stage ultrafiltration membrane module;
b) adding water and operating the tangential flow filtration system;
c) adjusting the inlet pressure and the outlet pressure; and
d) and determining the water flux of the system.
17. A method according to claim 16, wherein the multistage ultrafiltration membrane packages are cascaded multistage ultrafiltration membrane packages, preferably cascaded three stage ultrafiltration membrane packages.
18. The method of claim 16, further comprising an inlet pump in fluid communication with an inlet of the tangential flow filtration device, and a reflux control device in fluid communication with a reflux port of the tangential flow filtration device, preferably the reflux control device comprises a reflux port valve.
19. The method of claim 16, the tangential flow filtration system being the tangential flow filtration system of any of claims 1-8.
CN202010355618.5A 2020-04-29 2020-04-29 Tangential flow filtration system and method Pending CN113559712A (en)

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