Multi-module chromatography system

Abstract

A chromatography system comprising a mobile phase source A, an eluent phase source B, a sample source C, a waste tank D, a target tank E, a central control unit F and at least two chromatography units Gx, where each chromatography unit Gx comprises an inlet valve ax, a pump bx, a chromatography column cx having an inlet and an outlet, an outlet valve dx and a client control unit fx, where the inlet valve is in fluid communication with the pump via an inlet conduit, the outlet valve is in fluid communication with the pump via an outlet conduit, and the client control unit fx is in fluid communication with the pump via an outlet conduit. Wherein the pump is in fluid communication with an inlet of the chromatography column via a column conduit, where an outlet of the chromatography column is in fluid communication with an outlet of the chromatography unit via an outlet conduit, where the mobile phase source and the eluent phase source are in fluid communication with each inlet valve of each of the at least two chromatography units, preferably via separate flow paths, wherein the sample source is in fluid communication with the mobile phase source, preferably via a separate flow path, with each inlet valve of each of the at least two chromatography units, preferably via a sample-inlet joint, with each inlet conduit of each of the at least two chromatography units, each outlet of each of the at least two chromatography units is in fluid communication with a waste tank via a waste conduit, preferably via a sample-column joint sjx, in fluid communication with each column conduit of each of the at least two chromatography units, or a mixture thereof, where each outlet of each of the at least two chromatography units is in fluid communication with the waste tank via a sample-column joint sjx. Wherein each outlet of each of the at least two chromatography units is in fluid communication with a target tank via a target conduit, and wherein each client control unit of each of the at least two chromatography units cx is in communication with a central control unit F, wherein the outlet valve dx is preferably in fluid communication via a separate flow path with an inlet valve of a subsequent one of the at least two chromatographic units, preferably via a sample-inlet connection of the subsequent chromatographic unit with an inlet conduit of the subsequent chromatographic unit, the chromatographic unit is in fluid communication with a column conduit of a subsequent chromatographic unit, preferably via a sample-column junction of the subsequent chromatographic unit, or a mixture thereof, where the subsequent chromatographic unit is the first chromatographic unit G1 if the chromatographic unit is the last chromatographic unit Gx.

Description

Technical Field

[0001] This invention relates to multi-module chromatography systems. Background Technology

[0002] In biopharmaceutical manufacturing, such as the production of vaccines, antibodies, recombinant proteins, and gene therapy vectors, several chromatographic separation steps are typically required to remove various contaminants and impurities from the product. These separation steps significantly increase cost and processing time. Therefore, there is a general interest in increasing the yield and throughput of each step. Furthermore, many chromatographic steps used in downstream bioprocessing are performed discontinuously, i.e., in batch mode. Therefore, attempts are often made to perform downstream bioprocessing in a semi-continuous mode. Thus, it is beneficial to achieve maximum throughput for each batch step.

[0003] Typically, a single separation device, such as a column or membrane, is used in chromatographic processes. To address the problems mentioned above, multi-column continuous chromatography (MCC) has been developed, in which more than one separation device is interconnected. Typically, the separation devices are of the same type. Therefore, for example, in MCC mode, the chromatographic process involves the use of three columns with the same dimensions and the same stationary phase. If the columns are not identical, the theoretical calculations commonly used to design continuous chromatographic processes will be incorrect, and it will be difficult to design efficient and robust continuous chromatographic processes. The same argument applies if the feed concentration and flow rate change over time in unexpected ways.

[0004] In MCC, feed is applied to a first column and then transferred to one or more subsequent columns when the first column is near saturation. The first column is then eluted and regenerated to be reloaded during elution and regeneration of the subsequent(one or more) columns. Such processes are typically referred to as periodic countercurrent chromatography (PCC) or simulated moving bed (SMB) and offer considerable advantages for separations in biopharmaceuticals (see US7901581B1, US2013 / 0248451A1, US2013 / 0280788A1, and US7220356B1, the contents of which are incorporated herein by reference to the fullest extent permissible). PCC / SMB processes can significantly improve productivity, but the improvements achieved depend heavily on the design of the process and equipment.

[0005] Therefore, in MCC, in principle all columns can be run simultaneously, but the process steps change over time. The procedure can be repeated, allowing each column to be loaded, cleaned, and regenerated several times during the process. Compared to 'conventional' chromatography in continuous chromatography based on multiple identical columns, all steps, such as sample loading, stripping, clean-in-place (CIP), and rebalancing, occur simultaneously but individually on different columns.

[0006] Continuous chromatographic operations result in better utilization of chromatographic resins, i.e., stationary phases, reduced processing time, reduced resin volume required, and lower buffer requirements, all of which contribute to process economy. Summary of the Invention

[0007] However, while commercially available PCC systems are highly efficient, they are often very complex. Furthermore, such systems have certain limitations, such as being difficult to scale up or down, for example, in terms of the number of columns that can be used.

[0008] Therefore, the object of the present invention is to provide a modular, simple and easily expandable multi-column chromatography system that can be operated in the periodic countercurrent chromatography system in use.

[0009] It has now been surprisingly discovered that the aforementioned objective is achieved through a chromatographic system comprising a mobile phase origin, an elution phase origin, a sample origin, a sink, a target sink, a central control unit, and at least two chromatographic units. Each chromatographic unit includes an inlet valve, a pump, a column with an inlet and an outlet, an outlet valve, and a client control unit. The inlet valve is fluidly connected to the pump via an inlet conduit, the pump is fluidly connected to the inlet of the column via a column conduit, and the outlet of the column is fluidly connected to the outlet of the chromatographic unit via an outlet conduit. The mobile phase origin and the elution phase origin are preferably fluidly connected to each inlet valve of each of the at least two chromatographic units via separate flow paths. The sample origin is fluidly connected to the mobile phase origin, preferably via a separate flow path to each inlet valve of each of the at least two chromatographic units, and preferably via a sample-inlet connector to each inlet conduit of each of the at least two chromatographic units. Selectively, the column conduit of each of at least two chromatographic units is in fluid communication with the column conduit of each of the at least two chromatographic units via a sample-column connector, or a mixture thereof, wherein each outlet of each of the at least two chromatographic units is in fluid communication with the waste tank via a waste conduit, wherein each outlet of each of the at least two chromatographic units is in fluid communication with the target tank via a target conduit, and wherein each client control unit of each of the at least two chromatographic units communicates with the central control unit, wherein the outlet valve is preferably in fluid communication with the inlet valve of the subsequent chromatographic unit in the at least two chromatographic units via a separate flow path, preferably in fluid communication with the inlet conduit of the subsequent chromatographic unit via a sample-inlet connector of the subsequent chromatographic unit, preferably in fluid communication with the column conduit of the subsequent chromatographic unit via a sample-column connector of the subsequent chromatographic unit, or a mixture thereof, wherein if the chromatographic unit is the last chromatographic unit, then the subsequent chromatographic unit is the first chromatographic unit. Attached Figure Description

[0010] Figure 1 A schematic diagram of a chromatographic system according to the present invention is shown, which includes two chromatographic columns comprising a mobile phase (A), an eluent phase (B), and a sample phase (C).

[0011] Figure 2 A schematic diagram of a preferred embodiment of a chromatographic system comprising two chromatographic columns according to the present invention is shown, wherein the mobile phase (A) and the eluent phase (B) are provided from an active phase source (A1) and a reverse active phase source (B1), respectively.

[0012] Figure 3 A schematic diagram of a preferred embodiment of a chromatographic system according to the present invention is shown, the chromatographic system comprising three chromatographic columns including a mobile phase (A), an eluent phase (B), and a sample phase (C).

[0013] Figure 4 A schematic diagram of a preferred embodiment of a chromatographic system comprising three chromatographic columns according to the present invention is shown, wherein the mobile phase (A) and the eluent phase (B) are provided from the active phase source (A1) and the reverse active phase source (B1), respectively.

[0014] Figure 5 A schematic diagram of a preferred embodiment of a chromatographic system according to the present invention is shown, comprising a mobile phase (A), an eluent phase (B), and a sample phase (C), and further having additional communication between the column outlet and the inlet of a subsequent subsequent column (enabling post-load cleaning).

[0015] Figure 6 A schematic diagram of a preferred embodiment of a chromatographic system comprising three chromatographic columns according to the present invention is shown, wherein the mobile phase (A) and the eluent phase (B) are provided respectively from the active phase source (A1) and the reverse active phase source (B1), and further having additional communication between the column outlet and the inlet of a subsequent subsequent column (enabling post-load cleaning).

[0016] Figure 7 A schematic diagram of the steps is shown, according to Figure 1 The central control unit (F) of the chromatography system is configured to perform these steps to achieve periodic countercurrent chromatography in a binding / elution mode, without a post-load cleaning step.

[0017] Figure 8 A schematic diagram of the steps is shown, according to Figure 1 The central control unit (F) of the chromatography system is configured to perform these steps to achieve flow-mode periodic countercurrent chromatography without a post-load cleaning step.

[0018] Figure 9 A schematic diagram of the steps is shown, according to Figure 5The central control unit (F) of the chromatography system is configured to perform these steps to achieve periodic countercurrent chromatography in a binding / elution mode, including a post-loading cleaning step. Detailed Implementation

[0019] definition As the term "as used in this article" Chromatography system The term "chromatographic system" indicates a system comprising a pump, a first tubing, and a chromatographic column or column shortcut, wherein the first tubing provides fluid communication between the pump and the column or column shortcut. Preferably, the chromatographic system further comprises a second tubing and a detector and / or a fraction collector, wherein the second tubing provides fluid communication between the column or column shortcut and the detector and / or the fraction collector. The chromatographic system typically achieves input and output flows by connecting appropriate containers to the pump and / or detector via tubing. The chromatographic system may also include at least one valve and / or at least one bubble trap and / or at least one mixing chamber. Exemplary chromatographic systems are ÄKTA start, ÄKTA go, ÄKTA pure, ÄKTAavant, ÄKTV pilot, or ÄKTA ready, provided by Cytiva.

[0020] the term" unit "" indicates the apparatus included in the chromatographic system, wherein the different units of the chromatographic system are typically connected by test tubes. Preferably, the unit is selected from the list including: chromatographic column, column shortcut, detector, fraction collector, valve, bubble trap or mixing chamber.

[0021] As the term "as used in this article" detector "It can be any device suitable for detecting substances in a fluid passing through the apparatus. Preferably, the detector is selected from the following list: ultraviolet adsorption (UV) detector, visible light adsorption (VIS) detector, photodiode array (PDA) detector, refractive index detector, evaporative light scattering detector, multi-angle light scattering detector, mass spectrometer, conductivity detector, fluorescence detector, chemiluminescence detector, optical rotation detector, and electrochemical detector. Preferably, the detector is located downstream of the column."

[0022] As the term "as used in this article" chromatographic column "" indicates an apparatus for separating substances comprising a chromatographic matrix as a stationary phase.

[0023] the term" chromatographic matrix"" indicates the stationary phase in the chromatographic separation process. Preferably, the chromatographic matrix contains at least one active moiety that interacts with the target substance and / or impurities through one or more physical or chemical interactions. The chromatographic matrix can be fibrous, monolithic, membrane-like, or specific. Preferably, the chromatographic matrix is ​​a compound selected from the list including natural polymers and / or synthetic polymers, and more preferably a compound selected from the list including polysaccharides, polystyrene, polyacrylamide, polymethacrylates, or mixtures thereof.

[0024] As the term "as used in this article" beta-glucan "" indicates a macropolymer of dehydrated glucose with a molecular weight in the range of 3 kDa to 5 MDa. Dextran with a molecular weight greater than or equal to 2 MDa cannot penetrate the micropores of a chromatographic matrix, which comprises, and preferably is composed of, porous materials.

[0025] As the term "as used in this article" Active part "" indicates a molecule or substance that interacts with a target substance and / or impurities through one or more physical or chemical interactions. Preferably, the active moiety is selected from the list including: sulfopropyl, diethylaminoethyl, diethylaminopropyl, diethyl-(2-hydroxypropyl)aminoethyl, octylamine, N-benzyl-N-methylethanolamine, methanesulfonate, quaternary ammonium, carboxymethyl, alkyl, preferably octyl, hexyl, butyl, phenyl, biphenyl, pentafluorophenyl, phenylhexyl, ethyl, benzyl or isopropyl, silanol, peptide, nucleotide, protein, immunoglobulin-binding protein, preferably protein A, protein G, protein L, metal ion, preferably cations of Cu, Ni, Co, Zn, hypozinotriacetic acid, domain antibody or domain antibody fragment, CHT, polydeoxythymidine (e.g., 25-mer of dT), C4, C6, C8, C 10 C 12 C 14 C 16 Octadecyl carbon (C 18 C 20 C 22 C 24 Heparin, dextran sulfate, 2-mercaptopyridine, hydroxyapatite, fluorapatite, or combinations thereof.

[0026] As the term "as used in this article" Partial specific interactions"Affinity interaction," also known as an affinity interaction, refers to a specific interaction between a target substance and / or impurity and an interacting partner in the form of an active moiety. Preferably, the target molecule and / or impurity specifically binds to the active moiety. The moiety-specific interaction is preferably selected from: the interaction between an enzyme as a target substance and / or impurity and a substrate analog as a moiety; the interaction between an antigen as a target substance and / or impurity and an antibody as a moiety; the interaction between a polysaccharide as a target substance and / or impurity and a lectin as a moiety; the interaction between a complementary base sequence of a target substance and / or impurity and a nucleic acid as a moiety; the interaction between a hormone receptor as a target substance and / or impurity and a hormone as a moiety; and the interaction between a target substance and / or impurity and a hormone as a moiety. The interactions between the target substance and / or impurity biotin or biotin-binding substances (preferably biotin-binding proteins) and avidin as a component; the interactions between calmodulin binding partners as the target substance and / or impurity and calmodulin as a component; the interactions between the target substance and / or impurity glutathione S-transferase fusion substance (preferably glutathione S-transferase fusion protein) and glutathione as a component; the interactions between the target substance and / or impurity immunoglobulin and immunoglobulin-binding substances as a component (preferably immunoglobulin-binding proteins); and the interactions between the target substance and / or impurity polyhistidine fusion substance (preferably polyhistidine fusion protein) and metal cation-binding protein complex as a component.

[0027] As the term "as used in this article" fluid volume "" indicates a volume containing a fluid, and preferably containing the target species. Preferably, the fluid is a liquid. More preferably, the fluid is a liquid containing a solvent, preferably a liquid composed of a solvent. Even more preferably, the solvent is selected from aqueous buffer solutions, such as phosphates, acetates, citrates, or tris(hydroxymethyl)aminomethane, and mixtures thereof with other solvents (such as ethanol). The solvent may include additives, such as polymers (e.g., PEG and / or dextran), salts (e.g., ammonium sulfate, sodium chloride, etc.), cleaning agents, and / or stabilizers. Some affinity stationary phases (membranes or beads) may require more complex mixtures. For example, immobilized metal chelate affinity chromatography (IMAC) requires the addition of organic compounds such as imidazole.

[0028] As the term "as used in this article" Target types"" indicates the product to be separated. Typically, the target species is a substance naturally occurring in animal or plant tissues, produced by naturally occurring microorganisms or enzymes, and / or produced by modified (preferably genetically modified) microorganisms or enzymes. Therefore, the target species can be, but is not limited to, antibodies, proteins, viruses, organic matter, DNA molecules, RNA molecules, shorter nucleotide molecules, exosomes, cells (e.g., for cell therapy), or specific target molecules.

[0029] As the term "as used in this article" Raw material mixture "" indicates a mixture of various pollutants, solvents, and at least one target species. The mixture may be a natural product (such as animal or plant tissue) that has been in contact with at least one solvent, or a mixture from a bioreactor. Preferably, the feed mixture is a mixture from a bioreactor containing microorganisms such as yeast, bacteria and / or fungi, cells, viruses, tissues, the target species, and / or solvents.

[0030] As the term "as used in this article" Distribution Model "" indicates a chromatographic method in which impurities in the raw material mixture bind to the chromatographic matrix while the target species do not bind, thereby separating the target species from the diluent.

[0031] As the term "as used in this article" Binding / Elimination Mode "" indicates a chromatographic method in which the target species in the raw material mixture binds to the chromatographic matrix, while impurities do not bind. Impurities are washed from the column with a diluent, and the column is rinsed with an eluent that has removed the target species from the column, thereby separating the target species from the eluent.

[0032] As the term "as used in this article" Biological treatment unit "" indicates a unit that receives a fluid volume including a target species and typically includes contaminants, and outputs a fluid volume including the target species and typically includes fewer contaminants. Preferably, the biological treatment unit is selected from the list of the following: chromatographic devices including columns and membranes, filters, centrifuges, extraction devices, and two-phase separation devices.

[0033] As the term "as used in this article" Offline analysis Offline measurements indicate an analytical step in which samples are removed from a biological processing unit or transport line under aseptic conditions and analyzed in the laboratory after physical pretreatment (such as filtration and dilution). Preferably, preparation and processing require well-defined standard operating procedures (SOPs) and skilled personnel. Coupled with the complexity involved in manual processing, the main disadvantage of offline measurements is the time delay, which leads to a lower measurement frequency. Due to these issues, offline measurements are generally not considered true PAT (process analysis techniques) unless no other measurement possibilities exist (such as HPLC for product vibration or mass spectrometry for product quality). Offline laboratory measurements are typically used to monitor and validate the accuracy of in-line / online process analyzers.

[0034] As the term "as used in this article" Nearline analysis "" indicates an analytical step in which the sample is removed from the biological processing unit or transport line and analyzed manually or using an automated sampling device near the production process. Similar to offline measurements, aseptic conditions must be maintained for accurate results. Nearline measurements are most common for parameters that cannot be accurately measured in-line or online. Advantages of nearline measurements include reduced time delays (compared to offline) and the possibility of automated control.

[0035] As the term "as used in this article" Online analysis This indicates an analytical step in which the sample is transferred from the bioprocessing unit or transport line via a branch and can be returned to the bioprocessing process after analysis. The sample is automatically measured by process sensors in a bypass. The advantages of this method are its simple sterilization process and the direct acquisition of samples under fixed conditions. The implementation of this solution requires a specially designed or modified bioprocessing process, particularly the bioprocessing unit and / or transport line.

[0036] As the term "as used in this article" In-line analysis "" indicates an analytical step in which measurements are taken directly in the biological processing unit or transport line using process sensors. Preferably, the resulting measurements are sent to the control system in real time. Process parameters such as pH, ORP (oxidation-reduction potential), dissolved oxygen, dissolved CO2, temperature, and / or conductivity are typically measured via in-line analysis. Additionally, in-line analyses, such as UV / VIS absorbance / transmittance, fluorescence emission, and refractive index, can be used to measure measurements that allow for the detection of specific molecules.

[0037] As the term "as used in this article" Cleaning after loading "(PLW)" indicates a technique in the binding / elution mode where, after column loading is complete, the loaded column is washed with a diluent, i.e., the target species are not eluted from the column, thereby removing target species that do not bind to the chromatographic matrix. Preferably, the washing diluent is directed to a subsequent column to be loaded in the next cycle. Therefore, target species that have not yet bound to the chromatographic matrix that has undergone post-loading washing will undergo a binding process with the chromatographic matrix of the subsequent column. This ensures maximum binding of target species that were not bound after loading the first column.

[0038] As used in this specification and the appended claims, the singular forms “a” and “an” also include the corresponding plural forms, unless the context clearly specifies otherwise. In the context of this invention, the term “a” is used in conjunction with the plural forms. About "and" approximate "" indicates a precision range that a person skilled in the art would understand, in order to still ensure the technical effect of the feature in question. This term typically indicates a deviation from the indicated value of ±10%, preferably ±8%, more preferably ±5%, and even more preferably ±2%. It should be understood that the term "" include "and" Include "Not restrictive. For the purposes of this invention, the term " Composed of... "considered a term" include" Preferred embodiments. If a group is defined below as including at least a certain number of embodiments, this means it also includes a group preferably consisting only of these embodiments. Furthermore, the term " First " second " third "or" （a） " （b） " （c） " （d） The terms "..." are used to distinguish between similar elements and are not necessarily used to describe order or chronological order. It should be understood that such terms are interchangeable where appropriate, and embodiments of the invention described herein can operate in orders other than those described or shown herein. First " second " No. three "or" （a） " （b） " （c） " （d） In cases involving steps such as “i”, “K”, etc., relating to methods, uses, or measurements, there is no temporal or time interval consistency between the steps; that is, these steps may be performed simultaneously, or there may be time intervals of seconds, minutes, hours, days, weeks, months, or even years between these steps, unless otherwise indicated herein in the application set forth above or below. It should be understood that the invention is not limited to the specific methods, protocols, reagents, etc., described herein, as they may vary. It should also be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the invention, which will be limited only by the appended claims. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

[0039] As used in this article, the term " Not included " Not included "or not" means, in the context of the composition of the invention not containing a specific compound or a group of compounds that can be combined under common terms, that the composition does not contain more than 0.8% by weight of the compound or the group of compounds based on the total weight of the composition. Furthermore, preferably, the composition according to the invention does not contain more than 0.5% by weight of the compound or the group of compounds, and preferably the composition does not contain the compound or the group of compounds at all.

[0040] When referring to the composition and the weight percentage of its components, it should be understood that, according to the invention, the total amount of the components does not exceed 100% (±1% due to rounding).

[0041] Detailed description In various aspects, the present invention provides a multi-column modular chromatography system suitable for operation in continuous mode, preferably in periodic countercurrent continuous mode (PCC). This chromatography system is designed to combine existing single-column chromatographic units, such as ÄKTA start, ÄKTA go, ÄKTA pure, ÄKTA avant, ÄKTV pilot, or ÄKTAready provided by Cytiva, to form a multi-column modular chromatography system. This allows for modular design. Furthermore, it allows for easy exchange within the system not only of used columns but also entire chromatographic units. Finally, considering the chromatographic units, such as columns, the system is easily scalable.

[0042] The most general embodiment of the present invention is as follows: Figure 1 As shown in the figure. Therefore, the present invention provides a chromatography system comprising: Mobile phase source (A) Source of elution phase (B). Sample source (C). Waste trough (D) Target slot (E) Central control unit (F), and At least two chromatographic units (G x ), Each chromatographic unit (G) x )include: - Inlet valve (a x ), - Pump (b x ), - Chromatographic columns with inlet and outlet (c x ), - Outlet valve (d x ),as well as - Client control unit (f x ), Among them, the inlet valve (a) x ) via inlet conduit and pump (b x Fluid connectivity, Among them, pump (b) x) via column guide tube and chromatographic column (c x The inlet fluid is connected. Among them, the chromatographic column (c x The outlet of the chromatographic unit (G) is connected to the outlet conduit via the chromatographic unit. x ) exports (d x Fluid connectivity, The mobile phase source (A) and the eluent phase source (B) are preferably connected via separate flow paths to at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid connectivity, The sample source (C) and the mobile phase source (A) are in fluid communication, preferably via a separate flow path and at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and at least two chromatographic units (G x Each column conduit in each chromatographic unit is fluidly connected, or they are mixed. Among them, at least two chromatographic units (G x Each outlet of each chromatographic unit in the ) is fluidly connected to the waste tank (D) via a waste conduit. Among them, at least two chromatographic units (G x Each outlet of each chromatographic unit in the ) is fluidly connected to the target cell (E) via a target conduit, and Among them, at least two chromatographic units (G x Each client control unit (f) of each chromatographic unit in ) x ) communicates with the central control unit (F), Among them, the outlet valve (d) x Preferably via a separate flow path and at least two chromatographic units (G) x Subsequent chromatographic units (G) in ) x+1 ) inlet valve (a x+1 Fluid communication, preferably via a subsequent chromatographic unit (G) x+1 Sample - Inlet connector (si) x+1 ) and subsequent chromatographic units (G x+1 The inlet conduit is in fluid communication with the chromatographic unit (G), preferably via a subsequent chromatographic unit (G). x+1 Sample-column connector (sj) x+1 ) and subsequent chromatographic units (G x+1The columns and conduits are in fluid communication, or they are mixed. Where the chromatographic unit is the last chromatographic unit (G) x If ), then the subsequent chromatographic unit is the first chromatographic unit (G1).

[0043] Preferably, at least two chromatographic units (G x ( ) is a chromatographic module, such as ÄKTA start, ÄKTA go, ÄKTApure, ÄKTA avant, ÄKTV pilot, or ÄKTA ready. Preferably, at least two chromatographic units (G x ) chromatographic column (c x This includes a chromatographic matrix capable of adsorbing impurities to be separated from a target species or the target species itself. This has the advantage that the column can be loaded with the target species and subsequently eluted to obtain a clean target species. Therefore, the chromatographic matrix is ​​preferably suitable for affinity chromatography.

[0044] As described above, the sample source (C) and the mobile phase source (A) are in fluid communication, preferably via a separate flow path and at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and at least two chromatographic units (G x Each column conduit of each chromatographic unit in the system is in fluid communication with, or a mixture thereof. Therefore, the sample source (C) can be added to the system via a mobile phase source (A). Thus, in this embodiment, the sample is added directly to the mobile phase source (A). A disadvantage of this embodiment is that the sample will be distributed across a large amount of mobile phase, resulting in broadened chromatographic curves and reduced separation efficiency. Therefore, preferably, the sample source (C) is preferably connected to at least two chromatographic units (G) via a separate flow path. x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and at least two chromatographic units (G xEach column conduit in each chromatographic unit is fluidly connected, or they are mixed. Such an embodiment ensures that only a small volume of sample is added and reduces the broadening of the chromatographic curve. In particular, the sample source (C) and the inlet valve (a) x Fluid communication between the two valves has control that occurs only in a central control unit (i.e., the inlet valve). x The advantages of this method are that it utilizes sample sources (C) and at least two chromatographic units (G). x In ) each column conduit of each chromatographic unit is preferably connected via a sample-column connector (SJ). x Fluid connectivity of the sample-column joint has the advantage of minimizing the dead volume of the system because the sample-column joint (SJ) x ) is usually located as close as possible to the column (c x This reduces dilution and thus improves separation efficiency.

[0045] In a preferred embodiment of the invention, the waste conduit of the chromatography system will outlet (d x (A) is in fluid communication with the waste tank (D). Typically, during PCC chromatography, there will be a certain amount of diluent, eluent, and / or washing fluid, which contains little or no target species. These liquids are usually collected in the waste tank (D) for cleaning and reuse after the process is complete. Preferably, they are at least partially cleaned and separated during the process and reintroduced as (A) or (B).

[0046] Similarly, preferably, the target conduit of the chromatography system will exit (d x It is in fluid communication with the target tank (E).

[0047] Preferably, the mobile phase source (A) is a mobile phase reservoir, more preferably a mobile phase container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the mobile phase source (A) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0048] Preferably, the source of the eluent (B) is an eluent reservoir, more preferably an eluent container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the eluent source (B) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0049] Preferably, the sample source (C) is a sample reservoir, more preferably a sample container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the sample source (C) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0050] Preferably, the waste tank (D) is a waste reservoir, more preferably a waste container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the waste tank (D) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0051] Preferably, the target tank (E) is a target reservoir, more preferably a target container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the target tank (E) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0052] Typically, the reservoirs and containers of a chromatography system should be of appropriate size to accommodate the column volume and separation efficiency. Such appropriate dimensions reduce the risk of underperformance in the system.

[0053] The central control unit (F) can be any device for receiving, sending, and storing data, and for performing calculations using said data. Therefore, preferably, the central control unit (F) is a computer, preferably a computer connected to a network. However, the central control unit (F) can also be any other electronic device capable of performing the operations described herein. Therefore, the operation of the central control unit (F) can be implemented by hardware, for example, in electronic circuitry, or it can be implemented by software and configured to run on a computer. In a preferred embodiment of the invention, the central control unit (F) is a dedicated computer, i.e., a server, which is connected via a network to at least two chromatography units (G). x In an alternative preferred embodiment of the invention, the central control unit (F) may consist of at least two chromatographic units (G). x At least one of the following is provided. This embodiment has the advantage that the chromatography system does not necessarily require a dedicated computer.

[0054] Typically, at least two chromatographic units (G) x Each inlet (a) of each chromatographic unit in ) x The chromatographic unit (G) is configured to allow different or combined (preferably different) fluid communication between the mobile phase source (A), the eluent phase source (B), and / or the target source (C). More preferably, at least two chromatographic units (G) x Each inlet (a) of each chromatographic unit in ) x ) is configured to allow different or combined (preferably different) fluid communication between the mobile phase source (A), the eluent phase source (B), the target source (C), and / or the outlet valve (d) of the previous chromatographic unit. x-1 ) and inlet valve (a xThe inlet (ax) is preferably configured to allow fluid communication between the inlet conduit and one of the mobile phase source (A), the eluent phase source (B), the target source (C), or the outlet valve (d) of the previous chromatographic unit. x-1 ) and inlet valve (a x The fluid communication between at least two chromatographic units (G) is thus established. x Each inlet (a) of each chromatographic unit in ) x Preferably, it is a valve having at least four ports, and more preferably, it is an electrically actuated valve having at least four ports. More preferably, it has at least two chromatographic units (G... x Each inlet (a) of each chromatographic unit in ) x ( ) is a valve having at least 5 ports, preferably an electrically actuated valve having at least five ports.

[0055] Typically, at least two chromatographic units (G) x Each outlet (d) of each chromatographic unit in ) x ) is configured to allow different or combined (preferably different) fluid communication between the waste tank (D), the target tank (E), and / or the outlet valve (d x ) and subsequent chromatographic units (G x+1 The outlet valve (a) x+1 The fluid communication between at least two chromatographic units (G) is thus established. x Each outlet (d) of each chromatographic unit in ) x Preferably, it is a valve having at least 3 ports, and more preferably, it is an electrically actuated valve having at least 3 ports.

[0056] Preferably, in the chromatographic system according to the invention, at least two chromatographic units (G x ) includes at least three chromatographic units (G x This makes it possible to use a post-loading cleaning step.

[0057] More preferably, in the chromatographic system according to the invention, at least three chromatographic units (G x It includes at least four chromatographic units (G) x ).

[0058] In a preferred embodiment of the chromatographic system according to the invention, the mobile phase source (A) and the eluent phase source (B) are each provided free from the active phase source (A1) and the reverse active phase source (B1). Among them, the active phase source (A1) and the reverse active phase origin (B1) are associated with at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid connectivity, Among them, the sample source (C) and at least two chromatographic units (G) x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and at least two chromatographic units (G x Each column conduit in each chromatographic unit is fluidly connected, or they are mixed, and Among them, the inlet valve (a) x It is configured to provide a mixture of fluids supplied by an active phase source (A1), a reverse active phase source (B1), and / or a sample source (C).

[0059] The active phase source (A1) typically activates the interaction between the compound to be separated and the chromatographic matrix. Therefore, it depends on whether the chromatographic system is operating in binding / elution mode or flow-through mode. In binding / elution mode, the active phase source (A1) activates the interaction between the target species and the chromatographic matrix, while the reverse active source (B1) deactivates this binding. Similarly, in flow-through mode, the active phase source (A1) activates the interaction between the impurity to be removed and the chromatographic matrix, while the reverse active source (B1) deactivates this binding. Therefore, the ratio of these two sources can control the binding behavior of the target species and / or impurities in the column. Therefore, in this preferred embodiment, the inlet valve (a x It is typically configured to allow mixing of the active phase source (A1) and the reverse active phase source (B1). An advantage of this preferred embodiment is that the loading, elution, and optional post-loading washing steps can be performed with the same dilution components, but only in different proportions of (A1) and (B1).

[0060] Therefore, preferably, the inlet valve (a) x () is a mixing valve.

[0061] Preferably, the chromatographic system according to the invention includes a mixer (m) in the inlet conduit. x This ensures thorough mixing of the phases within the conduit.

[0062] Preferably, the active phase source (A1) is an active phase reservoir, more preferably an active phase container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the active phase source (A1) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0063] More preferably, the reverse active phase source (B1) is a reverse active phase reservoir, more preferably a reverse active phase container. Generally, any type of reservoir or container can be used. Preferably, the reservoir or container of the reverse active phase source (B1) is made of glass or a polymer, preferably polyethylene, polypropylene, or polyethylene terephthalate.

[0064] Preferably, in the chromatographic system according to a preferred embodiment of the present invention, the sample column connector is an injection valve (sj x Preferably, the injection valve (sj) x It is a 6-port rotary valve, preferably a 6-port electrically actuated valve.

[0065] Preferably, in the chromatographic system according to the invention, in at least two chromatographic units (G... x In each of the components, the column conduit includes a cleaning valve (sw) x ), of which at least two chromatographic units (G x Each cleaning valve (sw) in each chromatographic unit x The cleaning valve (sw) is in fluid communication with the waste tank (D) via a cleaning conduit. Preferably, the cleaning valve (sw) x It is a 3-port valve, preferably a 3-port electrically actuated valve.

[0066] Preferably, in the chromatographic system according to the invention, at least two chromatographic units (G x Each outlet conduit of each chromatographic unit in the ) includes at least one detector (g x The detector is selected from the list of the following: ultraviolet (UV) adsorption detector, visible light adsorption (VIS) detector, photodiode array (PDA) detector, refractive index detector, evaporative light scattering detector, multi-angle light scattering detector, mass spectrometer, conductivity detector, fluorescence detector, chemiluminescence detector, optical rotation detector, and electrochemical detector. Preferably, the detector (g x The detector is either an ultraviolet (UV) adsorption detector or a visible light adsorption (VIS) detector. More preferably, at least two chromatographic units (G... x Each outlet conduit of each chromatographic unit in the ) includes at least two detectors (g x Most preferably, at least two detectors (g) x This is a UV adsorption (UV) detector and a conductivity detector. The detector has the advantage of accurately and reliably detecting breakthroughs in target species and / or impurities within the column.

[0067] Preferably, the detector (g) x It can be part of online analysis, in-line analysis, or near-line analysis.

[0068] In a preferred embodiment of the invention, the chromatography system comprises at least two chromatographic units (G... x The client control unit of each chromatographic unit in the ) is configured to control the inlet valve (a x ), outlet valve (d x ) and pump (b x ), more preferably configured to control the inlet valve (a x ), outlet valve (d x ), pump (b x ), at least one detector (g x ) and injection valve (sj x ), more preferably configured to control the inlet valve (a x ), outlet valve (d x ), pump (b x ), cleaning valve (sw) x ), at least one detector (g x ) and injection valve (sj x Therefore, the term "control" specifically includes the action of actuating a valve, and more preferably includes opening and closing fluid communication with the valve.

[0069] Furthermore, in a preferred embodiment of the chromatography system of the present invention, the central control unit (F) is configured to be connected via a client control unit (f x In at least two chromatographic units (G) x The inlet valve (a) is controlled on each chromatographic unit in the ) x ), outlet valve (d x ) and pump (b x ), preferably configured to control the inlet valve (a x ), outlet valve (d x ), pump (b x ), at least one detector (g x ) and injection valve (sj x ), more preferably configured to control the inlet valve (a x ), outlet valve (d x ), pump (b x ), cleaning valve (sw) x ), at least one detector (g x ) and injection valve (sj x ).

[0070] Therefore, the control valve, in particular, includes the actuation of the actuating valve, more preferably including opening and closing fluid communication with the valve. Furthermore, the control detector includes retrieving measured values ​​from the detector. The control pump includes starting and stopping the pump and regulating the flow rate using the pump.

[0071] Preferably, in the chromatographic system according to the invention, the central control unit (F) is configured to enable the chromatographic system to perform periodic countercurrent chromatography (PCC). PCC can be performed in flow mode or in binding / elution mode, preferably in binding-elution mode. Furthermore, PCC can be performed with or without a post-loading cleaning step. If the central control unit (F) is configured to perform a post-loading cleaning step, then at least two chromatographic units (G) x It includes at least three chromatographic units.

[0072] Figure 1 The most general embodiment of the invention is shown, which includes at least two chromatographic units (G... x ). Figure 1 The central control unit (F) of the chromatographic system in the embodiment can be configured to operate the chromatographic system in binding / elution mode or in flow mode.

[0073] if Figure 1 In an embodiment, the central control unit (F) of the chromatographic system is configured to operate the chromatographic system in binding / elution mode, and is then configured to perform the following steps (see also...). Figure 7 ): In startup step a1, si1 and / or sj1 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d1 is actuated to establish fluid communication between d1 and a2, si2, and / or sj2, and a2, si1, and / or sj2 are actuated to establish fluid communication between d1 and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to establish fluid communication to the waste tank (D). Pumps b1 and b2 are then started, and detector g1 is monitored for breakthrough of the target species through the column (c1) of the first chromatographic unit (g1).

[0074] If a breakthrough is achieved, a1, si1, and / or sj1 are actuated in the first column elution step to close the fluid communication to the sample source (C) and establish fluid communication between the elution phase source (B) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close the fluid communication to a2, si2, and / or sj2 and establish fluid communication to the target tank (E). Furthermore, a2, si2, and / or sj2 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a2 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Then, detector g1 is monitored to indicate the end of elution in the column (c1) of the first chromatographic unit (G1).

[0075] If elution of the first column (c1) is complete and optional cleaning and recalibration are performed, a1, si1, and / or sj1 are actuated during the second column loading step to establish fluid communication between the outlet (d2) of the second chromatographic unit (G2) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Furthermore, d2 is actuated to establish fluid communication to a1, si1, and / or sj1 of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close fluid communication to the target tank (E) and establish fluid communication to the waste tank (D). Then, detector g2 is monitored for breakthrough of the target species through the column (c2) of the second chromatographic unit (G2).

[0076] If a breakthrough is achieved, a2, si2, and / or sj2 are actuated in the second column elution step to close the fluid communication to the sample source (C) and establish fluid communication between the elution phase source (B) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d2 is actuated to close the fluid communication with a1, si1, and / or sj1 and establish fluid communication to the target tank (E). Furthermore, a1, si1, and / or sj1 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Then, detector g2 is monitored to indicate the end of elution in the column (c2) of the first chromatographic unit (G2).

[0077] If elution of the second column (c2) is completed and optional cleaning and recalibration are performed, a2, si2, and / or sj2 are actuated during the first column loading step to establish fluid communication between the outlet (d1) of the first chromatographic unit (G1) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Furthermore, d1 is actuated to establish fluid communication to a2, si2, and / or sj2 of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to close fluid communication to the target tank (E) and establish fluid communication to the waste tank (D). Then, detector g1 is monitored for breakthrough of the target species through the column (c1) of the first chromatographic unit (g1).

[0078] If a breakthrough is achieved, the central control unit (F) is configured to also call the first column elution step again, followed by the second column loading step and the second column elution step, thereby achieving continuous chromatography.

[0079] If the process should be stopped, the central control unit (F) is configured to invoke the first or second column loading step, instead of redirecting the corresponding outlet to other columns, directly redirecting it to the waste tank (D) and the subsequent first or second column elution step.

[0080] According to Figure 2 In the embodiment where the mobile phase source (A) and the eluent phase source (B) are formed by a certain ratio of the active phase source (A1) and the reverse active phase source (B1), the central control unit is configured to implement [something related to] ... Figure 1 The same steps as in the embodiment, but a x si x and / or sj x Except for not being actuated to switch between (A1) and (B1) but to achieve a certain ratio required for the loading and elution steps. Therefore, a x si x and / or sj x The actuation is to achieve the proportions of (A1) and (B1) that allow the target species to be adsorbed onto the chromatographic matrix in the respective loading steps, and to achieve the proportions of (A1) and (B1) that allow the target species to be desorbed from the chromatographic matrix in the respective elution steps.

[0081] if Figure 1 The central control unit (F) of the chromatography system in the embodiment is configured to operate the chromatography system in flow mode, and is then configured to perform the following steps (see also...). Figure 8 ): In startup step a1, si1 and / or sj1 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d1 is actuated to establish fluid communication between d1 and a2, si2, and / or sj2, and a2, si2, and / or sj2 are actuated to establish fluid communication between d1 and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to establish fluid communication to the target tank (E). Pumps b1 and b2 are then started, and detector g1 is monitored for impurities breaking through the column (c1) of the first chromatographic unit (G1).

[0082] If a breakthrough is achieved, a1, si1, and / or sj1 are actuated during the first column cleaning step to close the fluid communication to the sample source (C) and establish fluid communication between the elution phase source (B) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close the fluid communication to a2, si2, and / or sj2 and establish fluid communication with the waste tank (D). Furthermore, a2, si2, and / or sj2 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a2 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Then, detector g1 is monitored to indicate the end of cleaning of the column (c1) of the first chromatographic unit (G1).

[0083] If the cleaning of the first column (c1) is completed and optional recalibration is performed, a1, si1, and / or sj1 are actuated in the second column separation step to establish fluid communication between the outlet (d2) of the second chromatographic unit (G2) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Furthermore, d2 is actuated to establish fluid communication to a1, si1, and / or sj1 of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close fluid communication to the waste tank (D) and establish fluid communication to the sample tank (E). Then, detector g2 is monitored for impurities breaking through the column (c2) of the second chromatographic unit (G2).

[0084] If a breakthrough is achieved, a2, si2, and / or sj2 are actuated in the second column cleaning step to close the fluid communication to the sample source (C) and establish fluid communication between the elution phase source (B) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d2 is actuated to close the fluid communication to a1, si1, and / or sj1 and establish fluid communication with the waste tank (D). Furthermore, a1, si1, and / or sj1 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). 。 Then, detector g2 is monitored to indicate the end of elution of column (c2) of the first chromatographic unit (G2).

[0085] If the cleaning of the second column (c2) is completed and optional recalibration is performed, a2, si2, and / or sj2 are actuated during the first column loading step to establish fluid communication between the outlet (d1) of the first chromatographic unit (G1) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Furthermore, d1 is actuated to establish fluid communication to a2, si2, and / or sj2 of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to close fluid communication to the waste tank (D) and establish fluid communication with the sample tank (E). Then, detector g1 is monitored for impurities breaking through the column (c1) of the first chromatographic unit (G1).

[0086] If a breakthrough is achieved, the central control unit (F) is configured to also call the first column cleaning step again, followed by the second column separation step and the second column cleaning step, thereby achieving continuous chromatography.

[0087] If the process should be stopped, the central control unit (F) is configured to invoke the first or second column separation step, instead of redirecting the corresponding outlet to other columns, directly redirecting it to the sample tank (E) and the subsequent first or second column cleaning step.

[0088] According to Figure 2 In the embodiment where the mobile phase source (A) and the eluent phase source (B) are formed by a certain ratio of the active phase source (A1) and the reverse active phase source (B1), the central control unit is configured to implement [something related to] ... Figure 1 The same steps as in the embodiment, but a x si x and / or sj xExcept for not being actuated to switch between (A1) and (B1) but to achieve a certain ratio required for the separation and cleaning steps. Therefore, a x si x and / or sj x The actuation is to achieve the proportions of (A1) and (B1) that allow impurities to be adsorbed onto the chromatographic matrix in the respective separation steps, and to achieve the proportions of (A1) and (B1) that allow impurities to be desorbed from the chromatographic matrix in the respective washing steps.

[0089] Figure 3-6 An embodiment of the present invention is shown, which includes at least three chromatographic units (G... x This allows for a post-loading wash step. The advantage of this step is that after column loading, at least a portion of the target species remains in the mobile phase on the column, and this portion is not adsorbed onto the chromatographic matrix. Since impurities are also present in the mobile phase, this mobile phase must be sent to waste before elution to ensure product purity. However, in the case of three columns in the system, the wash solution can be sent through a third, unloaded column, where the target species still present in the mobile phase can be adsorbed. This significantly improves the separation efficiency and yield of the chromatographic system. For post-loading wash to be possible, another interconnected column is necessary, as the wash solution from the post-wash step is sent to subsequent subsequent chromatographic units.

[0090] Therefore, preferably, in the chromatographic system according to the invention, the outlet valve (d) x ), preferably via a separate flow path and at least two chromatographic units (G x The subsequent chromatographic unit (G) x+2 ) inlet valve (a x+2 Fluid communication, preferably via a subsequent subsequent chromatographic unit (G). x+2 Sample - Inlet connector (si) x+2 ) and subsequent subsequent chromatographic units (G x+2 The inlet conduit is in fluid communication with the chromatographic unit (G), preferably via a subsequent chromatographic unit (G). x+2 Sample-column connector (sj) x+2 ) and subsequent subsequent chromatographic units (G x+2 The column and conduit are in fluid communication, or are mixed, where, if the chromatographic unit is the last chromatographic unit (G) x If the subsequent chromatographic unit is the second chromatographic unit (G2), and if the chromatographic unit is the penultimate chromatographic unit (G...), then the next chromatographic unit is the second chromatographic unit (G...). x-1 If the mobile phase is then sent to the first chromatographic unit (G1), then the subsequent subsequent chromatographic unit is the first chromatographic unit (G1). As described above, this feature enables the system to send the mobile phase to the subsequent subsequent chromatographic unit (G2). x+2 This is a requirement for cleaning after loading.

[0091] if Figure 5 In an embodiment, the central control unit (F) of the chromatography system is configured to operate the chromatography system in a binding / elution mode including post-load cleaning, and is then configured to perform the following steps (see also...). Figure 9 ): In startup step a1, si1 and / or sj1 are actuated to establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be established if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d1 is actuated to establish fluid communication between d1 and a2, si2, and / or sj2, and a2, si2, and / or sj2 are actuated to establish fluid communication between d1 and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to establish fluid communication to the waste tank (D). Pumps b1 and b2 are then started, and detector g1 is monitored for breakthrough of the target species through the column (c1) of the first chromatographic unit (G1).

[0092] If a breakthrough is achieved, in the first column cleaning step, a1, si1, and / or sj1 are actuated to close the fluid communication to the sample source (C) and establish fluid communication between the mobile phase source (A) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close the fluid communication to a2, si2, and / or sj2 and establish fluid communication to a3, si3, and / or sj3. Furthermore, a3, si3, and / or sj3 are actuated to establish fluid communication between outlet d1 and the inlet conduit, column conduit, and / or column of the third chromatographic unit (G3). Additionally, a2, si2, and / or sj2 are actuated to close the fluid communication to d1 and establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Alternatively, fluid communication to the mobile phase source (A) can be achieved if the sample source (C) is connected to the mobile phase source (A), or if a2 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d3 is actuated to achieve fluid communication to the waste tank (D). Then, detector g1 is monitored to indicate the end of cleaning of the column (c1) of the first chromatographic unit (G1).

[0093] If the end of elution is detected, a1, si1, and / or sj1 are actuated in the first column elution step to close the fluid communication to the mobile source (A) and establish fluid communication between the elution liquid source (B) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Simultaneously, outlet valve d1 is actuated to close the fluid communication to a3, si3, and / or sj3 and establish fluid communication to the target tank (E). Furthermore, outlet valve d2 is actuated to close the fluid communication to the waste tank (D) and establish fluid communication to a3, si3, and / or sj3. Additionally, a3, si3, and / or sj3 are actuated to establish fluid communication between outlet d2 and the inlet conduit, column conduit, and / or column of the third chromatographic unit (G3). Then, detector g1 is monitored to indicate the end of elution of the column (c1) of the first chromatographic unit (G1).

[0094] If a breakthrough is achieved, in the second column cleaning step, a2, si2, and / or sj2 are actuated to close the fluid communication to the sample source (C) and establish fluid communication between the mobile phase source (A) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to close the fluid communication to a3, si3, and / or sj3 and establish fluid communication to a1, si1, and / or sj1. Furthermore, a1, si1, and / or sj1 are actuated to establish fluid communication between outlet d2 and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Additionally, a3, si3, and / or sj3 are actuated to close the fluid communication to d2 and establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the third chromatographic unit (G3). Alternatively, fluid communication to the mobile phase source (A) can be achieved if the sample source (C) is connected to the mobile phase source (A), or if a3 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d1 is actuated to achieve fluid communication with the waste tank (D). Then, detector g2 is monitored to indicate the end of cleaning of the column (c2) of the second chromatographic unit (G2).

[0095] If the end of elution is detected, a2, si2, and / or sj2 are actuated in the second column elution step to close the fluid communication to the mobile source (A) and establish fluid communication between the elution liquid source (B) and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Simultaneously, outlet valve d2 is actuated to close the fluid communication to a1, si1, and / or sj1 and establish fluid communication with the target tank (E). Furthermore, outlet valve d3 is actuated to close the fluid communication to the waste tank (D) and establish fluid communication to a1, si1, and / or sj1. Additionally, a1, si1, and / or sj1 are actuated to establish fluid communication between outlet d3 and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Then, detector g1 is monitored to indicate the end of elution of the column (c1) of the first chromatographic unit (G1).

[0096] If a breakthrough is achieved, a3, si3, and / or sj3 are actuated in the third column cleaning step to close the fluid communication to the sample source (C) and establish fluid communication between the mobile phase source (A) and the inlet conduit, column conduit, and / or column of the third chromatographic unit (G3). Simultaneously, outlet valve d3 is actuated to close the fluid communication to a1, si1, and / or sj1 and establish fluid communication to a2, si2, and / or sj2. Furthermore, a2, si2, and / or sj2 are actuated to establish fluid communication between outlet d3 and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Additionally, a1, si1, and / or sj1 are actuated to close the fluid communication to d3 and establish fluid communication between the sample source (C) and the inlet conduit, column conduit, and / or column of the first chromatographic unit (G1). Alternatively, fluid communication to the mobile phase source (A) can be achieved if the sample source (C) is connected to the mobile phase source (A), or if a1 is a mixing valve that allows fluid communication between both the sample source (C) and the mobile phase source (A). Furthermore, d2 is actuated to achieve fluid communication to the waste tank (D). Then, detector g3 is monitored to indicate the end of column (c3) cleaning for the second chromatographic unit (G3).

[0097] If the end of elution is detected, a3, si3, and / or sj3 are actuated in the third column elution step to close the fluid communication to the mobile source (A) and establish fluid communication between the elution phase source (B) and the inlet conduit, column conduit, and / or column of the third chromatographic unit (G3). Simultaneously, outlet valve d3 is actuated to close the fluid communication to a2, si2, and / or sj2 and establish fluid communication to the target tank (E). Furthermore, outlet valve d1 is actuated to close the fluid communication to the waste tank (D) and establish fluid communication to a2, si2, and / or sj2. Additionally, a2, si2, and / or sj2 are actuated to establish fluid communication between outlet d1 and the inlet conduit, column conduit, and / or column of the second chromatographic unit (G2). Then, detector g3 is monitored to indicate the end of elution of the column (c3) of the first chromatographic unit (G3).

[0098] If the elution is completed, the central control unit (F) is configured to also call the first column cleaning step again, followed by the second column separation step and the second column cleaning step, thereby achieving continuous chromatography.

[0099] If the process should be stopped, the central control unit (F) is configured to invoke the first, second, or third column cleaning step, and instead of redirecting the corresponding outlet to other columns, redirect it directly to the waste tank (D).

[0100] According to Figure 6 In the embodiment where the mobile phase source (A) and the eluent phase source (B) are formed by a certain ratio of the active phase source (A1) and the reverse active phase source (B1), the central control unit is configured to implement [something related to] ... Figure 1 The same steps as in the embodiment, but a x si x and / or sj x Except for not being actuated to switch between (A1) and (B1) but to achieve a certain ratio required for the separation and cleaning steps. Therefore, a x si x and / or sj x The actuation is to achieve the proportions of (A1) and (B1) that allow impurities to be adsorbed onto the chromatographic matrix in the respective separation steps, and to achieve the proportions of (A1) and (B1) that allow impurities to be desorbed from the chromatographic matrix in the respective washing steps.

[0101] As shown in the figure, two or more self-contained chromatographic units Gx can be provided in a parallel configuration, each controllable via a corresponding client control unit (fx). This allows for easier system capacity expansion, as more chromatographic modules can be added to increase capacity as needed (e.g., in a plug-and-play configuration). Furthermore, replacement of any such chromatographic units is convenient, as they can be easily replaced without disrupting the operation of other chromatographic units. The chromatographic units are preferably provided as identical or substantially identical units.

[0102] Another advantage of the various embodiments of the present invention is that it provides a simpler system compared to conventional PCC chromatography, and this system (like PCC) is not limited to operation using only two columns. For example, there is a need to reduce the number of valves and other complex and expensive equipment (such as additional UV monitors and pumps). Furthermore, it enables simplified and reliable control strategies, rather than the more complex control strategies that may be involved in conventional PCC and SMB setups.

[0103] Furthermore, various embodiments of the invention can also operate continuously, for example, automatically. In one test run, such a system (in this case using columns overloaded by the Delta-UV method) ran continuously for 48 hours without encountering any problems.

[0104] The chromatographic unit can also be provided as a single-use (SU) unit. This allows embodiments of the invention to provide a "PCC-like" chromatographic system with single-use technology. Currently, there are no known single-use PCC or SMB systems due to the high cost of the existing technologies used.

[0105] Various other advantages of the present invention will also be apparent to those skilled in the art.

Claims

1. A chromatographic system, comprising: Mobile phase source (A) Elution liquid phase source (B). Sample source (C). Waste trough (D) Target slot (E) Central control unit (F), and At least two chromatographic units (G x ), Each chromatographic unit (G) x )include: - Inlet valve (a x ), - Pump (b x ), - Chromatographic columns with inlet and outlet (c x ), - Outlet valve (d x ),as well as - Client control unit (f x ), Among them, the inlet valve (a) x ) via the inlet conduit to the pump (b) x Fluid connectivity, Wherein, the pump (b) x ) via column guide tube and the chromatographic column (c x The inlet fluid is in communication with the ) Wherein, the chromatographic column (c x The outlet of the chromatographic unit (G) is connected to the chromatographic unit (G) via an outlet conduit. x The outlet (d) x Fluid connectivity, The mobile phase source (A) and the eluent phase source (B) are preferably connected to the at least two chromatographic units (G) via separate flow paths. x Each inlet valve (a) of each chromatographic unit in ) x Fluid connectivity, The sample source (C) is in fluid communication with the mobile phase source (A), and preferably via a separate flow path with the at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and the at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and the at least two chromatographic units (G x Each column conduit in each chromatographic unit is fluidly connected, or they are mixed. Among them, the at least two chromatographic units (G x Each outlet of each chromatographic unit in the ) is in fluid communication with the waste tank (D) via a waste conduit. Among them, the at least two chromatographic units (G x Each outlet of each chromatographic unit in the ) is in fluid communication with the target cell (E) via a target conduit, and Among them, the at least two chromatographic units (G x Each client control unit (f) of each chromatographic unit in ) x ) communicates with the central control unit (F), Among them, the outlet valve (d) x Preferably via a separate flow path with the at least two chromatographic units (G) x Subsequent chromatographic units (G) in ) x+1 The inlet valve (a) of the above x+1 Fluid communication, preferably via the subsequent chromatographic unit (G) x+1 The sample-inlet connector (si) of the sample x+1 ) and the subsequent chromatographic unit (G x+1 The inlet conduit is in fluid communication with the subsequent chromatographic unit (G), preferably via the subsequent chromatographic unit (G). x+1 The sample-column connector (sj) x+1 ) and the subsequent chromatographic unit (G x+1 The column conduits described herein are in fluid communication, or are a mixture thereof. Wherein, if the chromatographic unit is the last chromatographic unit (G) x If ), then the subsequent chromatographic unit is the first chromatographic unit (G1).

2. The chromatographic system according to claim 1, wherein, The outlet valve (d) x Preferably via a separate flow path with the at least two chromatographic units (G) x Subsequent chromatographic units (G) in ) x+2 The inlet valve (a) of the above x+2 Fluid communication, preferably via the subsequent subsequent chromatographic unit (G) x+2 The sample-inlet connector (si) of the sample x+2 ) and the subsequent subsequent chromatographic unit (G x+2 The inlet conduit is in fluid communication with the subsequent chromatographic unit (G), preferably via the subsequent chromatographic unit (G). x+2 The sample-column connector (sj) x+2 ) and the subsequent subsequent chromatographic unit (G x+2 The column conduits described herein are in fluid communication, or are a mixture thereof. Wherein, if the chromatographic unit is the last chromatographic unit (G) x If the subsequent chromatographic unit is the second chromatographic unit (G2), and if the chromatographic unit is the penultimate chromatographic unit (G...), then the subsequent chromatographic unit is the second chromatographic unit (G...). x-1 If the subsequent chromatographic unit is the first chromatographic unit (G1), then the subsequent chromatographic unit is the first chromatographic unit (G1).

3. The chromatographic system according to claim 1 or 2, wherein, The waste conduit will lead to the outlet (d) x It is in fluid communication with the waste tank (D).

4. The chromatographic system according to any one of the preceding claims, wherein, The target catheter will lead to the outlet (d) x It is in fluid communication with the target tank (E).

5. The chromatographic system according to any one of the preceding claims, wherein, The at least two chromatographic units (G) x Each inlet of each chromatographic unit in the ) is a valve with at least 4 ports, preferably an electrically actuated valve with at least 4 ports.

6. The chromatographic system according to any one of the preceding claims, wherein, The at least two chromatographic units (G) x Each outlet of each chromatographic unit in the ) is a valve with at least 3 ports, preferably an electrically actuated valve with at least 3 ports.

7. The chromatographic system according to any one of the preceding claims, wherein, The mobile phase source (A) and the elution phase source (B) are each provided by the active phase source (A1) and the reverse active phase source (B1), respectively. Wherein, the active phase source (A1) and the reverse active phase source (B1) are related to the at least two chromatographic units (G). x Each inlet valve (a) of each chromatographic unit in ) x Fluid connectivity, Wherein, the sample source (C) and the at least two chromatographic units (G) x Each inlet valve (a) of each chromatographic unit in ) x Fluid communication, preferably via a sample-inlet connector (si) x ) and the at least two chromatographic units (G x Each inlet conduit of each chromatographic unit in the ) is fluidly connected, preferably via a sample-column connector (sj x ) and the at least two chromatographic units (G x Each column conduit in each chromatographic unit is fluidly connected, or their mixing is possible, and Among them, the inlet valve (a) x ) is configured to provide a mixture of fluids supplied by the active phase source (A1), the reverse active phase source (B1), and / or the sample source (C), preferably wherein the inlet valve (a x () is a mixing valve.

8. The chromatographic system according to claim 7, wherein, The inlet conduit includes a mixer (m x ).

9. The chromatographic system according to any one of the preceding claims, wherein, The at least two chromatographic units (G) x Each outlet conduit of each chromatographic unit in the ) includes at least one detector (g x The detectors are selected from the following list: ultraviolet light adsorption (UV) detectors, visible light adsorption (VIS) detectors, photodiode array (PDA) detectors, refractive index detectors, evaporative light scattering detectors, multi-angle light scattering detectors, mass spectrometers, conductivity detectors, fluorescence detectors, chemiluminescence detectors, optical rotation detectors, and electrochemical detectors.

10. The chromatographic system according to any one of the preceding claims, wherein, The client control unit is configured to control the inlet valve (a) x The outlet valve (d) x ) and the pump (b x ), preferably configured to control the inlet valve (a x The outlet valve (d) x The pump (b) x ), the at least one detector (g x ) and the injection valve (sj x More preferably, it is configured to control the inlet valve (a) x The outlet valve (d) x The pump (b) x The cleaning valve (sw) x ), the at least one detector (g x ) and the injection valve (sj x ).

11. The chromatographic system according to claim 10, wherein, The central control unit (F) is configured to communicate via the client control unit (f) x ) in at least two chromatographic units (G x The inlet valve (a) is controlled on each chromatographic unit in the ) x The outlet valve (d) x ) and the pump (b x ), preferably configured to control the inlet valve (a x The outlet valve (d) x The pump (b) x ), the at least one detector (g x ) and the injection valve (sj x More preferably, it is configured to control the inlet valve (a) x The outlet valve (d) x The pump (b) x The cleaning valve (sw) x ), the at least one detector (g x ) and the injection valve (sj x ).

12. The chromatographic system according to claim 11, wherein, The central control unit (F) is configured to communicate via the client control unit (f x ) actuates the inlet valve (a x The outlet valve (d) x ) and the pump (b x ), preferably configured to control the inlet valve (a x The outlet valve (d) x The pump (b) x ), the at least one detector (g x ) and the injection valve (sj x More preferably, it is configured to control the inlet valve (a) x The outlet valve (d) x The pump (b) x The cleaning valve (sw) x ), the at least one detector (g x ) and the injection valve (sj x This enables the chromatographic system to perform periodic countercurrent chromatography (PCC).

13. The chromatographic system according to claim 11 or 12, wherein, The central control unit (F) is configured to perform periodic countercurrent chromatography in flow mode or in binding-elution mode, preferably in binding-elution mode.

14. The chromatographic system according to any one of the preceding claims, wherein, The at least two chromatographic units (G) x ) includes at least three chromatographic units (G x ).

15. The chromatographic system according to claim 14, wherein, The central control unit (F) is configured to perform at least one post-load cleaning step.

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