REGULATED PROCESS FOR THE SEPARATION OF A MIXTURE

Description

FIELD OF THE INVENTION

[0001] The present invention relates to a chromatographic method for separating a mixture. TECHNICAL BACKGROUND

[0002] Chromatography is a separation technique based on the difference in the distribution of compounds in a mixture between a mobile phase and a stationary phase. Compounds are separated by percolating a mobile phase consisting of a liquid, gaseous, or supercritical solvent through a device (called a column or cell) filled with the stationary phase. A separation occurs when all or some of the compounds have a different percolation rate from each other. This method is used as an analytical technique to identify and quantify the compounds in a mixture. It can also be used as a purification technique.

[0003] Depending on the needs, different chromatographic processes are used to achieve the purification of molecules. Some are described in the document Large Scale Adsorption and Chromatography, PC Wankat, CRC press, Boca-Raton, 1986, the document Adsorption and Ion Exchange , MD Le Van et al., Perry's Chemical Engineers' Handbook 7th edition, Section 16, Mac-Graw-Hill, N-1, 1997 and the document Choice and optimization of operating mode in industrial chromatography , RM Nicoud et al., Proceeding of the 9th International Symposium on Preparative and Industrial Chromatography, PREP 92, April 1992, Nancy, pp. 205-220. These processes can be classified according to several criteria: the process can be either batch or continuous, the systems can have one or more columns, the composition of the eluent can be isocratic or a gradient can be used.

[0004] Several methods for regulating these processes have been described, notably in documents FR 2699917, US 5,457,260, FR 2762793, DE 19842550, US 2005 / 0107895 and the article Optimal operation of simulated moving bed chromatographic processes by means of simple feedback control, H. Schramm et al., Journal of Chromatography A, 2003, vol. 1006, p.3-13.

[0005] Document EP 0878222 describes an SMB device comprising at least one concentration detector located in the fluid circulation passage, means located in the outlet lines for measuring the concentration of a compound in the extract and / or in the raffinate and an operation controller determining the control conditions for the operation of the device on the basis of the concentration data from the concentration detector and the means located on the outlet lines.

[0006] WO 2007 / 101944 describes a chromatographic method in which a detector determines the history of a variable at a node of the chromatography device. In this history, a characteristic point located in an area between a successive raffinate collection and an extract collection surrounding the injection of the mixture to be separated, a characteristic point located on the adsorption front, a characteristic point located on the desorption front are detected, and the positions of these points are compared to target positions. The quantity of mobile phase in the areas bearing these characteristic points is then adjusted to make the position of the characteristic points coincide with their target position.

[0007] This process allows in particular to maximize the quantity of mixture treated but has the disadvantage of not allowing the control of the quantity of mobile phase injected. However, in certain applications, it is useful to be able to control as closely as possible the volume of mobile phase used for the separation of the compounds of a mixture, in order in particular to control the consumption of eluent or the quantity of energy necessary for the concentration of the purified products. In addition, if the volume of eluent injected is too low, the viscosity of the collected fractions can become too high, which can be disadvantageous for their subsequent treatment, in particular by passage through an evaporator.

[0008] There is therefore a need to provide a method for regulating a chromatographic separation, in which the operating parameters are optimized to meet purity or yield constraints while allowing precise control of the volume of mobile phase injected, and not requiring the use of a separation model or a complex algorithm. SUMMARY OF THE INVENTION

[0009] The invention relates firstly to a method for separating a mixture in a system comprising a plurality of chromatography columns, the method comprising successively, cyclically, in a given part of the system: a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract and a step of injecting mobile phase; the system comprising zones 1, 2, 3 and 4, zone 1 being located between a mobile phase injection line and an extract collection line, zone 2 being located between said extract collection line and an injection line of the mixture to be separated, zone 3 being located between said injection line of the mixture to be separated and a raffinate collection line, and zone 4 being located between said raffinate collection line and said mobile phase injection line; the method further comprising: determining, at a node of the system, the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; detecting on said history a single characteristic point in the portion of the history corresponding to zone 1, zone 4, and the interface between these two zones; comparing the position of the characteristic point with respect to a target position;adjusting the carrier volume of zone 1 and / or the carrier volume of zone 4, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position; the volume of the mobile phase injected per cycle being kept greater than or equal to a minimum limit and / or less than or equal to a maximum limit.;

[0010] According to embodiments, the volume of mobile phase injected per cycle is maintained at a constant value.

[0011] According to embodiments, the target position of the characteristic point is defined relative to the position of the mobile phase injection step and / or the position of the extract collection step and / or the position of the raffinate collection step.

[0012] According to embodiments, the variable representative of the concentration of one or more species contained in the mixture to be separated is chosen from a rotatory power, an absorbance, an emission of spectroscopic radiation, a refractive index, a density, a conductivity, a pH and combinations thereof.

[0013] According to embodiments, the characteristic point is chosen from a local minimum of the history, a point of the history having a determined value and a barycenter of two points of the history corresponding to values defined absolutely or relatively.

[0014] According to embodiments, the method further comprises the steps of: measuring the purity and / or yield of at least one collected fraction selected from the extract and the raffinate; comparing the measured purity and / or yield with a target purity and / or yield.

[0015] According to embodiments, the method comprises the steps of: measurement of the purity and / or yield of the raffinate and the purity and / or yield of the extract; comparison of the measured purity and / or yield of each fraction with a target purity and / or yield respectively.

[0016] According to embodiments, the method further comprises a step of modifying the volume of mixture to be separated injected per cycle as a function of the difference between the purity(ies) and / or the yield(s) measured and the purity(ies) and / or the target yield(s).

[0017] According to embodiments, the method further comprises a step of modifying the volume of mobile phase injected per cycle as a function of the difference between the measured purity(ies) and / or yield(s) and the target purity(ies) and / or yield(s).

[0018] According to embodiments, the method further comprises a step of jointly modifying the volume of mixture injected per cycle and the volume of mobile phase injected per cycle as a function of the difference between the purity(ies) and / or the yield(s) measured and the purity(ies) and / or the target yield(s).

[0019] According to embodiments, the method further comprises a step of defining the target position as a function of the difference between the measured purity(ies) and / or yield(s) and the target purity(ies) and / or yield(s).

[0020] According to embodiments, the method further comprises: the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; the detection on the history of another characteristic point in the portion of the history corresponding to zone 2, zone 3, and the interface between these two zones; the comparison of the position of the other characteristic point with respect to a target position; the adjustment of the carrier volume of zone 2 and / or the carrier volume of zone 3 modifying the position of the other characteristic point to bring it closer to its target position.

[0021] According to embodiments, the injection of the mixture to be separated is a continuous flow.

[0022] According to embodiments, the method is a simulated moving bed process.

[0023] According to embodiments, the injection of the mixture to be separated is discontinuous.

[0024] According to embodiments, the method is a sequential simulated moving bed process.

[0025] According to embodiments, the mixture to be separated comprises one or more monosaccharides, preferably glucose and fructose, and the extract and the raffinate are enriched in different monosaccharides.

[0026] According to embodiments, the volume of the mobile phase injected per cycle is maintained greater than or equal to a minimum limit and / or less than or equal to a maximum limit, and the method comprises: the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; the process being such that: as long as the volume of the mobile phase injected per cycle is greater than or equal to said minimum limit, or less than or equal to said maximum limit, the method comprises: ▪ the detection on the history of a first characteristic point in the portion of the history corresponding to zone 1; ▪ the detection on the history of a second characteristic point in the portion of the history corresponding to zone 4; ▪ the comparison of the position of each of the characteristic points with a respective target position; ▪ the adjustment of the carrier volume of zone 1 and the carrier volume of zone 4, respectively modifying the position of the first characteristic point and the second characteristic point, to bring them closer to their respective target positions;- when the volume of the mobile phase injected per cycle reaches the maximum limit or the minimum limit, the method comprises: ▪ the detection on said history of a single characteristic point in the portion of the history corresponding to zone 1, zone 4, and the interface between these two zones; ▪ the comparison of the position of the characteristic point with respect to a target position; ▪ the adjustment of the carrier volume of zone 1 and / or the carrier volume of zone 4, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position. ;

[0027] The invention also relates to a computer program comprising program code instructions for executing the steps of the method as described above when said program is executed on a computer.

[0028] The invention also relates to a computer-readable storage medium on which a computer program as described above is recorded.

[0029] The invention also relates to a system comprising a processor coupled to a memory on which is recorded a computer program as described above.

[0030] The present invention overcomes the drawbacks of the prior art.

[0031] According to a first aspect, it is based on regulation of a chromatographic separation system making it possible to optimize operating parameters such as flow rates or periods, having one or more advantageous properties among the following properties: the process makes it possible to meet the purity and / or yield requirements of the collected fractions, the process allows adaptation to variations in chromatographic conditions by supporting the inertias intrinsic to the chromatographic systems and / or by supporting possible analysis delays, in particular purity and yield analysis, the process makes it possible to satisfy constraints on the volume of mobile phase injected, whether it is a maximum volume not to be exceeded or a minimum volume to be respected.

[0032] The invention is based on: the use of a characteristic point called " low concentration »located on the history of a representative variable between the start of an extract collection step and the end of the next raffinate collection step, the position of which is brought back to a target position by an adjustment of the carrier volume of said characteristic point; in conjunction with the fixing of a minimum limit and / or a maximum limit for the volume of mobile phase injected, per cycle.

[0033] In document WO 2007 / 101944, two characteristic points of the history, respectively dedicated to adsorption and desorption, are controlled (see figures 22 and 23 and the corresponding passage of the description). According to the invention, a single characteristic point is controlled instead of these two characteristic points. Compared to the teaching of document WO 2007 / 101944, this control of the position of a single characteristic point, less perfectly defining the position of the two adsorption and desorption fronts, makes it possible in return to control the quantity of mobile phase injected in each cycle precisely and in particular to guarantee a controlled evolution between a maximum and / or minimum quantity of mobile phase injected.

[0034] The present disclosure also describes, according to a second aspect not representing the invention, a method which is based on a purity (and / or yield) measurement of one or more collected fractions not requiring an off-line collection device dedicated to analyzing the purity of the collected fraction. The method according to this second aspect has one or more advantageous properties among the following: the method allows rapid and simple obtaining of purity measurement of the collected fraction, the method allows savings in terms of cleaning, the method allows simplification of the device necessary to carry out purity measurements and also allows increasing the frequency of measurements, since the steps of cyclical collections of representative sample, cleaning and preparation of the device are eliminated.

[0035] This is achieved by using at least one online detector and determining the histories of at least two variables representative of the concentration of at least two species contained in the mixture to be separated by means of this detector. A detector such as a spectrometer can measure several representative variables. This purity measurement can be used for plant monitoring or combined with the use of different control algorithms.

[0036] The present disclosure also describes, according to a third aspect not representing the invention, a method which is based on a measurement of purity (and / or yield) of one or more collected fractions not requiring an off-line collection device dedicated to the analysis of the purity of the collected fraction. The method according to this third aspect has one or more advantages among the following: the method is suitable for the use of slow detectors and can be implemented when the use of fast detectors is not possible, the method can be implemented in conventional installations in which a storage tank preceding another system such as a concentration means is present, the method does not require a device which needs to be purged and cleaned between each purity analysis.

[0037] This is achieved by measuring the concentrations of the collected fraction species in an intermediate tank present on the collected fraction outlet line. This purity measurement device can be used for plant monitoring or combined with the use of different control algorithms.

[0038] In certain embodiments of the methods disclosed herein, the first aspect is implemented in conjunction with the second aspect.

[0039] In certain embodiments of the methods disclosed herein, the first aspect is implemented in conjunction with the third aspect. BRIEF DESCRIPTION OF THE FIGURES

[0040] The following figures are obtained from accumulation chromatographic processes, and more particularly from SMB or SSMB processes. There figure 1represents a density history on which three examples of low concentration characteristic points are shown. The x-axis corresponds to a volume index representing the volume observed at the observation node relative to the total volume observed at the observation node over a cycle. The y-axis corresponds to the density. The figure 2 represents a rotational power history on which three examples of low concentration characteristic points are shown. The x-axis corresponds to a volume index representing the volume observed at the observation node relative to the total volume observed at the observation node over a cycle. The y-axis corresponds to the polarity. The Figures 3 and 4 represent a density history obtained during the implementation of a method according to the invention in which a maximum eluent volume constraint ( figure 3 ) or minimal ( figure 4 )was imposed. This history shows the position of the eluent injection (PM), the characteristic adsorption point (symbol "O" on the right side of the figure), its target position (symbol "+" on the right side of the figure), the characteristic desorption point (symbol "O" on the left side of the figure), its target position (symbol "+" on the left side of the figure), the characteristic low concentration point (symbol "◊") and its target position (symbol "x"). The abscissa axis corresponds to the ratio of volume observed at the observation node to the total volume observed at the observation node over a cycle. The ordinate axis corresponds to the density. The extract collection is noted E, the raffinate collection is noted R. figures 5 .A, 5.B and 5.Crepresent the concentration graphs (in g / L) of the species to be separated as a function of the ratio of the advancement volume to the installed resin volume (bed volume (BV)), obtained by a chromatographic simulation model in accordance with the examples section below, with different injected quantities. The species monitored are a mixture of glucose polymers (DPn (degree of polymerization) greater than 1) (curve consisting of diamonds), glucose (curve consisting of squares) and fructose (curve consisting of triangles). The Figure 5 .A represents the concentration graph obtained during a charge pulse, the Figure 5 .B represents the concentration graph obtained during saturation at high concentration and the Figure 5 .C represents the concentration graph obtained during saturation at low concentration. The figures 6 , 7, 8represent the evolution of the position of the characteristic desorption points ( Figure 6 ), adsorption ( Figure 7 ), low concentration ( Figure 8 )measured and the respective target position of each of these characteristic points (in volume ratio to the cycle volume, relative to the average injection position of the eluent) as a function of the number of cycles when implementing the processes described in Example 1. The target position of the characteristic point is represented by the symbol "o", the position of the characteristic point measured when implementing the process regulated using the two characteristic points of adsorption and desorption and without eluent volume constraint is represented by the symbol "+" and the position of the characteristic point measured when implementing the process regulated using the low concentration characteristic point and with a maximum eluent volume constraint is represented by the symbol "x". figures 9 And 13represent the eluent consumption per cycle (in bed volume (BV)) as a function of the number of cycles when implementing the processes described in Examples 1 and 2. The value of the maximum eluent volume constraint ( Figure 9 ) or minimal ( Figure 13 ) imposed is represented by the symbol "o", the actual eluent consumption per cycle when implementing the regulated process using the two adsorption and desorption characteristic points and without eluent volume constraint is represented by the symbol "+" and the actual eluent consumption per cycle when implementing the regulated process using the low concentration characteristic point and with an eluent volume constraint represented by the symbol "x". figures 10 , 11 and 12 present the same data (represented in the same way) as the figures 6 , 7 and 8, but obtained when implementing the methods described in Example 2, the symbols "x" representing the position of the characteristic point measured when implementing a process regulated using the low concentration characteristic point and with a minimum eluent volume constraint. The figures 14 , 15 , 16 , 17 , 18 And 21 represent the evolution of purity and yield (y-axis on the left, in percentage) and the evolution of the volumes of eluent and mixture injection (y-axis on the right, in volume ratio to column volume (bed volume - BV) as a function of the number of cycles when implementing the process described in example 3 ( figure 14 ), example 4 ( figure 15 ), example 5 ( figures 16 , 17 and 18 )and example 7 ( figure 21 ). The target purity is represented by the gray “□” symbol (curve 1), the target yield is represented by the gray “o” symbol (curve 2), the measured purity is represented by the gray “▪” symbol (curve 3), the measured yield is represented by the gray “•” symbol (curve 4), the volume of eluent injected per cycle is represented by the black “x” symbol (curve 5) and the volume of mixture to be separated injected per cycle is represented by the “+” symbol (curve 6). figures 19 and 20 represent the evolution of the optical rotation of glucose ( figure 19 ) and fructose ( figure 20 ) depending on temperature and glucose concentration ( figure 19 ) and fructose ( figure 20 )obtained during the calibration of the polarimeter in example 6. The x-axis corresponds to the concentration of the species (glucose or fructose) in g / L. The axis of the ordinates corresponds to the rotational power (or rotation angle). Rotational powers are measured at a temperature of 20°C (symbol " ◊ "), 35°C (symbol "▪"), 45°C (symbol " ▲ "), 55°C (symbol "x") and 65°C (symbol " "). DESCRIPTION OF EMBODIMENTS OF THE INVENTION

[0041] The invention is now described in more detail and in a non-limiting manner in the following description. General presentation of the chromatographic separation process

[0042] The three aspects described in the present disclosure relate to a method for separating a mixture in a system comprising a set of several chromatography columns containing a stationary phase, the method successively comprising, in a cyclic manner, in a given part of the system: a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract and a step of injecting mobile phase.

[0043] The above steps occur in succession in this part of the system. The relevant part of the system is preferably located between the output of one column and the input of the next column. Alternatively, the relevant part of the system may include a column or part of a column.

[0044] At any given time, one or more of the above steps may be simultaneously implemented in one or more parts of the system. For example, all of these steps may be simultaneously implemented in respective parts of the system.

[0045] By "mixture to separate ", means a mixture of species (or compounds, including molecules) containing at least two species, for example at least one species of interest and at least one impurity. The mixture to be separated may be binary, when it is composed of two species, or complex, when it is composed of more than two species. The mixture to be separated may be diluted in a liquid phase, preferably the mobile phase used in the chromatographic process.

[0046] According to embodiments, the mixture to be separated comprises one or more species chosen from: a monosaccharide sugar, for example glucose, fructose, deoxyribose, ribose, arabinose, xylose, lyxose, ribulose, xylulose, allose, altrose, galactose, gulose, idose, mannose, talose, psicose, sorbose or tagatose, and / or a polysaccharide sugar, for example a galacto-oligosaccharide, a fructo-oligosaccharide or a wood hydrolysate, and / or proteins, and / or amino acids, and / or organic acids such as citric acid, and / or mineral salts, and / or ionized species, and / or alcohols and / or glycols, and / or organic acids from natural or enzymatic or fermentative media.

[0047] This list is not exhaustive; the invention in its entirety can be carried out on any chemical species to be separated.

[0048] In some embodiments, the mixture to be separated comprises one or more monosaccharides. Preferably, the extract and the raffinate are enriched in different monosaccharides. Advantageously, the monosaccharide comprises 5 or 6 carbon atoms. Preferably, the monosaccharide is selected from glucose, fructose, deoxyribose, ribose, arabinose, xylose, lyxose, ribulose, xylulose, allose, altrose, galactose, gulose, idose, mannose, talose, psicose, sorbose, tagatose and a mixture thereof. In some particularly advantageous embodiments, the mixture to be separated comprises glucose and fructose.

[0049] The method according to the first aspect, corresponding to the invention, is particularly advantageous in such a case, since the consumption of the eluent represents a significant cost in the separation of these monosaccharides.

[0050] In this text, the terms "mixture to separate"", “load”, “mixture to be treated”, “product to be purified” And "initial mixture » mean the same thing. The terms " mobile phase » And "eluent" » mean the same thing in this text.

[0051] By "raffinate", we mean a fraction enriched in species less retained by the stationary phase. In the case of an initial binary mixture, this is the fraction enriched in the least retained species.

[0052] By " extract » we mean a fraction enriched in species more retained by the stationary phase. In the case of an initial binary mixture, this is the fraction enriched in the most retained species.

[0053] The chromatographic columns are preferably arranged in series and in a closed loop, with one outlet of one column connected to a subsequent column inlet, and the outlet of the last column connected to the inlet of the first column.

[0054] Columns can also be called " cells »chromatography. They can be used in a carousel system, arranged side by side, or arranged one above the other in one or two towers to limit floor space.

[0055] The columns may contain a liquid or solid stationary phase. The eluent may be a gaseous, liquid, or even supercritical fluid. Injection lines for the mixture to be separated and the eluent are provided at the inlet of the various columns, and extract and raffinate collection lines are provided at the outlet of the columns. Preferably, the injection and collection lines are connected to connection lines between two successive columns.

[0056] In certain advantageous embodiments, the system also comprises members for sequencing the injection and collection lines. In particular, the sequencing of these injection and collection lines takes place over an operating cycle of the system. In the present text, a " operating cycle » Or " cycle » refers to the time after which the injection and collection lines have been sequenced until they return to their initial position in the system. At the end of a cycle, the system is again in its initial configuration. A cycle generally has as many periods as there are columns in the separation loop. Thus, the cycle of a process implemented on an 8-column system is made up of 8 periods.

[0057] Moving the collection lines (extract and raffinate) and injection lines (feed and mobile phase) in the system is also referred to as line switching in this description.

[0058] Four zones can be defined in the system (especially when the process implemented is an SMB process, as described below): zone 1 located between the eluent injection line and the extract collection line, zone 2 located between the extract collection line and the mixture injection line to be separated, zone 3 located between the mixture injection line to be separated and the raffinate collection line, and zone 4 located between the raffinate collection line and the eluent injection line.

[0059] The method according to the invention is advantageously a periodic accumulation chromatographic method.

[0060] By "process has accumulation”, means a chromatographic process in which the injection of the mixture to be separated is intercalated or added to a non-zero concentration profile passing from the outlet to the inlet of a column.

[0061] Examples of such accumulation processes are the SMB process, the VariCol process, the Powerfeed process, the ModiCon process, the iSMB process or the SSMB process.

[0062] The Simulated Moving Bed (or SMB) process is a continuous multi-column process, with the injection of the mixture to be separated being carried out over the entire cycle.

[0063] The SMB process is a four-zone SMB process. In this case, the system comprises a set of columns connected in series and in a closed loop, the outlet of one column being connected to the inlet of the next column. The system comprises at least one injection line for the mixture to be separated, a collection line for a fraction enriched in species that is poorly retained by the stationary phase (the raffinate), an injection line for an eluent, and a collection line for a fraction enriched in species that is more retained by the stationary phase (the extract). The injection lines (for the mixture to be separated and the eluent) and the fraction collection lines move periodically and synchronously (synchronous sequencing) within the loop in the direction of the flow of the fluid circulating through the loop.The time between two shifts of all the injection and collection lines of a column corresponds to a period; at the end of a cycle all the points have returned to their initial position, the system having a cyclical operation. A cycle has as many periods as there are columns.

[0064] The process according to the invention may be a process with continuous injection of the mixture to be separated (i.e. a process in which the injection of the mixture to be separated is a continuous flow). The injection of the mixture to be separated is then carried out throughout the duration of the cycle. The process according to the invention may also be a process with quasi-continuous injection of the mixture to be separated. In certain particularly advantageous embodiments, the process according to the invention is an SMB process, preferably a four-zone SMB process.

[0065] Alternatively, the method according to the invention may be a method in which the injection of the mixture to be separated is discontinuous. In these methods, the injection of the mixture to be separated is not carried out over the entire cycle but for a total duration of less than one cycle. As a method with discontinuous injection of the mixture to be separated, the iSMB method may be mentioned. (“Improved Simulated Moving Bed” or improved SMB in French), described in documents EP 0342629 and US 5,064,539. In this process, in one step the system operates in a closed loop, without injection or collection of product. We can also cite the SSMB process (“Sequential Simulated Moving Bed” or sequential SMB in French) whose multi-column sequential is for example described in document WO 2015 / 104464. Preferably, the method according to the invention, when it involves discontinuous injection of the mixture to be separated, is an SSMB method. Regulation process

[0066] The method according to the first aspect, which corresponds to the invention, comprises: the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; the detection on said history of a single characteristic point between the start of a step of collecting the extract and the end of the following step of collecting the raffinate; the comparison of the position of the characteristic point with respect to a target position; the adjustment of the carrier volume of the characteristic point, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position; the volume of the mobile phase injected per cycle being kept above a minimum limit and / or below a maximum limit.

[0067] By "knot of the system" or "observation node", we mean a physical point of the chromatography system which can be freely chosen.

[0068] In some embodiments, the observation node is located between the output of one column and the input of the next column in the system.

[0069] We call "historical", the state or evolution of a variable representative of the concentration of one or more species contained in the mixture to be separated moving in the system, this state being considered for a duration or time determined at the observation node. Thus, we can measure the evolution of a variable representative of the concentration of one or more species contained in the mixture to be separated flowing at the observation node. A history can be represented: as a change over time, for example in a non-limiting manner expressed in gross time, or in time elapsed in relation to the start of the cycle or reduced to the total cycle time; as a change over time of the integration of a liquid flow rate, preferably the flow rate flowing at the observation node, for example in a non-limiting manner expressed in gross volume, or in volume elapsed in relation to the start of the cycle or reduced to another volume (volume of a column or total cycle volume), this method being particularly interesting in the case where the flow rates are variable during the period or the cycle; as a more general change in an indicator of progress in the cycle.

[0070] The history can thus be temporal, volumetric or dependent on a parameter setting of the process cycle.

[0071] A history is different from a concentration profile. We call it "concentration profile"the concentration state of the fractions of the fluid flowing in the system, this state being considered at a given moment over the entire system. Document FR 2699917 cited at the beginning of this description describes steps for reconstructing a concentration profile.

[0072] The duration during which the state of the variable is determined, i.e. during which the history is determined, is for example an operating cycle. At the end of the cycle, the history can be reset and started again. The duration can also be shorter than an operating cycle.

[0073] As indicated above, the abscissa of a history can be expressed in a non-limiting way in different units: raw time: the axis then starts from 0 and ends at the actual end of the cycle time; reduced time, defined by the raw time divided by the cycle time: in this case, the history axis is always between 0 and 1; a cycle progress index: this is the generalization of the raw time divided by the cycle time; this is useful in the case where there are scheduled stops of elution flow rates whose duration can be variable; a volume index corresponding to the volume observed at the system node divided by the total volume observed at the observation node.

[0074] In advantageous embodiments, the chromatographic system comprises a detector, and possibly a plurality of detectors, positioned at the node of the system where the history of the variable representative of the concentration of one or more species contained in the mixture to be separated is obtained.

[0075] The detector can be, for example, a densimeter, a polarimeter, a conductivity meter, a refractometer, an infrared, near infrared, Raman or UV / visible spectrometer or an online nuclear magnetic resonance device.

[0076] The detectors can be located on the system, i.e. on the system lines themselves; for this purpose, it is possible to envisage that the fluid circulates through the detectors. This is advantageous for low fluid flow rates in the system. For higher flow rates, it may be preferable to move the detectors away from the lines. For this purpose, bypasses are made on which the detectors are placed.

[0077] The method according to the invention comprises a step of determining, at a node of the system, the history of a variable representative of the concentration of one or more species contained in the mixture to be separated. Thus, the state of a variable representative of the concentration of one or more species is determined, by means of one or more detectors as described above.

[0078] This variable may be the concentration of one or more species of the mixture if the detectors allow it. In other embodiments, during this step, the variable representing the concentration of one or more species is not the concentration itself or the purity itself of the fractions. This has the advantage of regulating the operation more quickly than if a concentration or a purity had to be measured (such a measurement can be complex and therefore time-consuming). Furthermore, since it is not necessary to measure the purity or the concentration, it is not necessary to calibrate the detector and the method according to the invention can thus tolerate a drift of the detector(s) used.

[0079] It is not necessary to physically create a history, for example by viewing or printing the history. Direct regulation without recording or viewing may be sufficient. Preferably, the history determination step is carried out in a single node, which limits the use of detectors and avoids having to consider the synchronization of the detectors, the synchronization can also evolve over time in the case, for example, of a change in state of one of the columns of the system.

[0080] The variables representing the concentration of one or more species contained in the mixture to be separated are preferably variables that can be obtained quickly and directly, unlike, for example, the concentration which often requires prior calibration. Thus, as an example of a specific variable, we can cite: the optical rotation, obtained for example by the signal returned by a polarimetric detector, usable in the case where the species of the mixture to be separated are optically active, for example enantiomers; the absorbance or emission of spectroscopic radiation, obtained for example by the signal returned by a spectroscopic detector, for example UV / visible or infrared radiation, usable when the species of the mixture to be separated are natural or synthetic molecules having detectable chemical groups, in particular biomolecules such as proteins or peptides; the refractive index, the density, the conductivity or the pH, obtained for example by the signal returned by detectors measuring such physical quantities, usable for example in the case where the mixture to be separated contains sugars, ionic species, acids or bases;the combination of several specific variables cited above, one or more variables obtained from other detectors.;

[0081] The concentration variables given above as examples are variables for which a history can be easily obtained. The histories of these variables can be obtained in real time, which makes the process efficient. The histories reflect the evolution of concentrations at the observation node, in the form of a signal that can easily be obtained with detector monitoring.

[0082] The separation method according to the invention also comprises a step of detecting a characteristic point on the history, called " characteristic point of low concentration »in the present description. This step makes it possible, from the determination of the variable representative of the concentration, to define a point revealing the separation process. According to certain embodiments, the characteristic point is not a precise value of the variable but is revealing of a phenomenon circulating at the observation node. The characteristic point is revealing of a relative behavior of the species circulating at the observation node.

[0083] The observation node is positioned at the outlet of a column of the system. During the cycle, the lines (raffinate collection, injection of the mixture to be separated, extract collection and mobile phase injection) and the zones move from one column to another and said lines and zones are therefore located at precise times next to (or in the vicinity) of the observation node, that is to say between the same successive columns as the observation node. On the history, it is possible to define times and durations corresponding to the passage of the lines (raffinate collection, injection of the mixture to be separated, extract collection and mobile phase injection) in the vicinity of the observation node.It is then possible to identify on the history the intervals or average points of the positions (also called average positions) of collection of the raffinate, injection of the mixture to be separated, collection of the extract and injection of mobile phase as well as the zones that these intervals and average points determine.

[0084] According to the invention, the characteristic point of low concentration is located between the start of an extract collection step and the end of the following raffinate collection step.

[0085] On the history, said start of the extract collection step and said end of the following raffinate collection step frame a mobile phase injection step. The method according to the invention is a four-zone method, as described above, and the characteristic point of low concentration is located either in zone 1, or in zone 4, or at the interface between these two zones (i.e., at the time corresponding to the characteristic point, the observation node is located either in zone 1 of the system, or in zone 4, or at the interface between these two zones, i.e. at the average injection position of the eluent).

[0086] In embodiments, the detection of the characteristic point on the history occurs between the end of an extract collection step and the start of the next raffinate collection step.

[0087] In embodiments, the detection of the characteristic point takes place in a portion of the history in which the density and / or the total concentration of the species is less than or equal to 25% of the maximum total density or, respectively, concentration of the mixture to be separated. The detection of the low concentration characteristic point corresponds to the detection of the overall position of the assembly formed by the adsorption front and the desorption front on the history, whereas in document WO 2007 / 101944, the positions of the two fronts are individually detected and used to carry out the control of the positions of the adsorption and desorption fronts.

[0088] By "adsorption front",we mean an increase in concentration observed at the column outlet, in particular when the concentration increases from a low value to a value close to the maximum concentration detected over a cycle. The concentration can be the concentration of a species in the mixture to be separated or the concentration of all the species present.

[0089] By " desorption front”, we mean a decrease in concentration observed at the column outlet, in particular when the concentration decreases from a value close to the maximum concentration detected on a cycle to a low value. The concentration can be the concentration of a species of the mixture to be separated or the concentration of all the species present.

[0090] The characteristic point of low concentration can be, for example: the minimum of the history, that is to say the minimum signal point as detected between the collection of the extract and the collection of the raffinate, or a local minimum; this point can be measured by a minimum signal of a densimeter, a UV detector, but also by the minimum of the absolute value of the signal of a polarimeter; the point of the history having a determined value, for example the zero value, in particular when the variable representing the concentration is the optical rotation and the species to be separated have optical rotations of opposite signs; a point calculated as an intermediate between two points corresponding to predetermined values on the history; the predetermined values can for example correspond to a threshold of the signal of a densimeter or a UV detector, but also to a threshold of the absolute value of the signal of a polarimeter;the characteristic point may be, for example, a barycenter and in particular the isobarycenter (temporal or volumetric) of two points in the history corresponding to predetermined values; a point calculated as an intermediary between two points corresponding to values defined relatively on the history; these values may in particular correspond to respective fractions of the values reached at local maxima or minima of the history; thus the characteristic point may be a barycenter and in particular the isobarycenter (temporal or volumetric) between two points corresponding to such defined values, and in particular between two points which are respectively a characteristic point of the adsorption front and a characteristic point of the desorption front; the advantage of this method is that these two characteristic points can be calculated according to two different detectors. ;

[0091] THE Figures 1 and 2present examples of low concentration characteristic points detected from a history. The figure 1corresponds to a density history (black line), obtained by a densimeter, between the collection of the extract (denoted E) and that of the raffinate (denoted R). Three examples of low concentration characteristic points are presented. A first example of a low concentration characteristic point, denoted 1, is the point corresponding to the minimum density. A second example is the point denoted 2 whose abscissa is the isobarycenter of the abscissas of the two points of the history corresponding to a predetermined density value of 1.05.A third example is point 3 whose abscissa is the isobarycenter of the abscissas of two points obtained by relative threshold at 20%: the first point is defined by the abscissa such that the density is at 20% between the minimum density observed and the density measured at the start of extract collection; the second point is defined reciprocally by the abscissa such that the density is at 20% between the minimum density observed and the density measured at the end of raffinate collection. The average position of the mobile phase injection is noted PM.

[0092] There figure 2corresponds to a history of optical rotation, obtained by a polarimeter, between the collection of the extract (denoted E) and that of the raffinate (denoted R). Three examples of low concentration characteristic points are presented. A first example of a low concentration characteristic point is the point denoted 1 where the measured optical rotation is zero. A second example is the point denoted 2 whose abscissa is the isobarycenter of the abscissas of the two points corresponding to a threshold value of the absolute optical rotation value. A third example is the point denoted 3 whose abscissa is the isobarycenter of the abscissas of the point corresponding to a optical rotation equal to 50% of the maximum optical rotation and of the point corresponding to a optical rotation equal to 50% of the minimum optical rotation. The position of the mobile phase injection is denoted PM.

[0093] The position of the characteristic point is data used to stabilize the purities and yields of the system.

[0094] As indicated above, an advantage of the method according to the invention is that the history can be determined over a period of less than one cycle. Indeed, it is sufficient to detect the characteristic point to then move on to the next step of the regulation. Preferably, however, the determination step is carried out over a complete cycle. Furthermore, the steps of determining the history and detecting the characteristic point can be implemented at a frequency corresponding to an integer number of cycles (every n cycles, n being greater than or equal to 1). The more frequently the steps of determining the history and detecting the characteristic point are implemented, the more precise the adjustment of the system operation.

[0095] The method according to the invention also comprises a step of comparing the position of the characteristic point with a target position.

[0096] The detection of the characteristic point can be likened to the determination of the time (or the elapsed volume, the latter corresponding to the integration of the flow rate over time) where the characteristic point appears at the observation node during the duration of the history, for example a cycle; the target position then corresponds to the time (or the volume) where the characteristic point should appear at the observation node to allow operation under the desired conditions.

[0097] On a history corresponding to the evolution over time of a variable representative of the concentration of one or more species contained in the mixture to be separated, the step of comparing the position of the characteristic point with a target position consists of comparing the abscissa of the characteristic point with a predetermined target abscissa. This makes it possible to determine whether a disturbance has occurred in the system. In a properly adjusted system, preferably without disturbance and in steady state, the position of the characteristic point coincides with the target position. If the system is not disturbed, the characteristic point appears at an observation node at approximately the same time in each cycle. The difference between the position of the characteristic point and the target position can then correspond to a difference in time spent at the observation node.

[0098] The target position of the low concentration characteristic point can be defined absolutely in the cycle or relative to the mobile phase injection step, and / or relative to a fraction, raffinate and / or extract collection step.

[0099] The method according to the invention also comprises a step of adjusting the carrier volume of the characteristic point, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position, if a difference appears between the position of the characteristic point and the target position. In other words, a difference between the position of the characteristic point and the target position corresponds to a deviation or a disturbance in the operation of the system which can be compensated for by adjusting only the carrier volume of the characteristic point.

[0100] By " volume carrying a concentration front”,for the purposes of the present invention, the carrying volume of a front on which the characteristic point is located or in the vicinity of which the characteristic point is located is understood.

[0101] In the four-zone process according to the invention, as described above, this may be the carrier volume of zone 1 (or desorption zone), or the carrier volume of zone 4 (or adsorption zone). For “load-bearing volume of an area”,we mean the volume circulating in the zone in question between two line switches. This volume is the product of the fluid flow rate circulating in the zone and the period, it can also be the integration over time of the flow rate between two line switches when the flow rate is not constant during the period. When the mobile phase volume of zone 1 is modified, the position of the desorption front present in zone 1 is also modified. Similarly, the volume of zone 4 carries the adsorption front and a modification of the volume of zone 4 will modify the position of the adsorption front.

[0102] By " load-bearing volume of the characteristic point”, within the meaning of the present invention, a volume whose variation modifies the position of the characteristic point is understood. By observing the Figures 1 and 2 ,a change in the volume of zone 1 will change the position of the front between the start of extract collection and the average eluent injection point and will change the position of the characteristic point of the desorption front. And conversely, a change in the volume of zone 4 will change the position of the front between the average eluent injection point and the end of raffinate collection, thus changing the position of the characteristic point of the adsorption front.

[0103] A modification of the carrier volume of the characteristic point can be carried out by modifying one or more injection and / or collection rates; and / or by modifying the duration between line switchings; and / or by modifying the duration of one or more injections and / or collections. Preferably, the modification of the carrier volume of the characteristic point is carried out by modifying one or more injection and / or collection rates or the duration between line switchings.

[0104] To change the carrying volume of the characteristic point, for example, the flow rate in zone 1 or the flow rate in zone 4 can be changed. The flow rate will be increased, for example, if the position of the characteristic point is located after the target position (i.e., if the abscissa of the characteristic point is located after the abscissa predetermined as the target position) or reduced if the position of the characteristic point is located before the target position.

[0105] Preferably, the carrier volume of the characteristic point is adjusted without changing the mobile phase injection volume. The mobile phase volume injected per cycle can then remain constant during use of the method. This is particularly useful when a mobile phase volume constraint is set. Thus, when the mobile phase volume injected per cycle reaches the set extreme limit (whether it is a minimum or maximum limit), this volume remains constant at this limit. Changing the carrier volume of the adsorption zone (zone 4), for example by changing the raffinate collection flow rate, then amounts to changing the carrier volume of the desorption zone (zone 1) by the same amount, and vice versa.

[0106] Thus, adjusting the carrying volume of the characteristic point allows the passage of species in the area whose carrying volume is modified to vary. The characteristic point may be ahead of the target point (i.e., the point whose abscissa is the target position). By reducing the carrying volume of the characteristic point, the circulation of species can be slowed down and the characteristic point delayed so as to bring it closer to the target point. Conversely, if the characteristic point is behind the target point, the carrying volume of the characteristic point is increased to accelerate the circulation of species and accelerate the characteristic point so as to bring it closer to the target point. If the characteristic point is stabilized at the target point, this means that there is no adjustment to be made.It is also possible to consider providing a difference threshold between the position of the characteristic point and the target position; the adjustment is then carried out if the observed difference exceeds the threshold.

[0107] A regulation as described above can be for example a PID regulation (Proportional, Integral, Derivative regulation) acting on the carrier volume of the characteristic point.

[0108] In a four-zone process, it is necessary to distinguish between the terms “load-bearing volume of an area” (also called "volume of an area" Or "mobile phase volume of an area"), "mobile phase volume" And “volume of mobile phase injected”.

[0109] Thus (and as explained above), the volume of zone 1 corresponds to the volume of the mobile phase which passes over a period of time through the column(s) specific to zone 1, located between the eluent injection line and the extract collection line. In a similar manner, a volume can be defined for each zone of the system (for zone 2, for zone 3 and for zone 4). As indicated above, the modification of a zone volume is preferably carried out either by a modification of the flow rate, or by a modification of the time between two switchings of the lines which frame said zone. This can modify the volume of mobile phase injected or the volume of mobile phase of the collections. For example, a variation of the volume of zone 4 while leaving constant the volumes of zone 3 and zone 1 is equivalent to a variation of the raffinate collection volume and the volume of mobile phase injected.

[0110] The term of "mobile phase volume"is a generic term for the mobile phase passing through a column.

[0111] On the other hand, the " injected mobile phase volume " Or " volume of mobile phase injected per cycle » corresponds to the volume of fresh eluent injected between zone 4 and zone 1 (in a four-zone process). It is this volume that the invention aims to control.

[0112] The control method described in WO 2007 / 101944 allows the individual control of the mobile phase volumes of zones 1 and 4, which are the volumes carrying the characteristic points of zones 1 and 4 (desorption and adsorption characteristic points). This does not allow the precise control of the mobile phase volume injected per cycle.

[0113] In the method according to the invention, when a constraint on the volume of mobile phase injected per cycle is encountered (i.e. when a maximum and / or minimum limit is reached), the regulation is carried out by making the same correction to the volume of zone 1 and to the volume of zone 4 (the method being a four-zone method). The regulation thus relates to a quantity called " carrier volume of the low concentration point » which may be, in the case of the four-zone process according to the invention, the volume of zone 1 or the volume of zone 4 or a combination of the two. In such a case, a variation made to the volume of zone 1 and zone 4 will modify the collection volume of the raffinate and / or the extract but not the volume of mobile phase injected.

[0114] The advantage of the method according to the invention is that it allows both rapid reaction to disturbances in the system and control of the quantity of mobile phase injected at each operating cycle, unlike the methods described in the prior art. Generally speaking, methods based on the analysis of the composition of collected fractions do not allow rapid reaction to disturbances whose effects take several cycles to stabilize. In addition, in certain cases, the time taken to analyze and obtain purity values is long. The time taken to analyze the compositions is all the longer since it is preferable to sample the fractions of the chromatographic system over an entire cycle, these samples then being analyzed. Thus, the purity results can only be obtained with at least one cycle delay and with a periodicity greater than one or two cycles, which makes regulation and determination of the setting to be applied more difficult.There is therefore a delay in the reactivity of storage systems following changes in operating parameters or following disturbances, to which an analysis delay may be added.

[0115] The present method makes it possible to react as soon as a change in the history is observed. The regulation implemented to restore the system is then rapid. In addition, the method according to the invention also makes it possible, by controlling a characteristic point between the start of an extract collection step and the end of the following raffinate collection step, to control the volume of mobile phase used by the chromatographic process, unlike processes regulated by controlling two characteristic points located on the adsorption front and the desorption front.

[0116] There figure 3shows a history obtained when using a method according to the invention in which a constraint of maximum volume of eluent injected per cycle has been set. The position of the low concentration characteristic point coincides with its target position. In this example, the position of the low concentration characteristic point corresponds to the position of the eluent injection. It is noted that the adsorption and desorption characteristic points (as described in WO 2007 / 101944) have not reached their target position and that deviations persist between the adsorption and desorption points and their respective target position.

[0117] There figure 4shows a history obtained when using a method according to the invention in which a minimum volume constraint of eluent injected per cycle has been set. The position of the low concentration characteristic point coincides with its target position. In this example also, the position of the low concentration characteristic point corresponds to the position of the eluent injection. It can be seen that in this case also the adsorption and desorption characteristic points have not reached their target position and that deviations persist between the adsorption and desorption points and their respective target position.

[0118] This regulation based on the characteristic point of low concentration and not on the characteristic points of adsorption and desorption allows compliance with the imposed eluent volume extrema.

[0119] According to certain embodiments, the method further comprises a step of measuring the purity of at least one collected fraction. In particular, the method comprises a step of measuring the purity of the extract and / or the raffinate.

[0120] The purity of the fraction(s) is then compared with a target purity, i.e. a predetermined purity that one wishes to achieve.

[0121] It is also possible, as an alternative to measuring the purity(ies) or in addition to these measurements, to measure a quantity of a target species in a collected fraction and determine its yield linked to the presence of target species in the other collected fraction(s). Thus, the method according to the invention may comprise a step of measuring the yield of a target species in the extract and / or raffinate and a step of comparing the measured yield(s) with a target yield respectively. In the present invention, it is possible to use purity and / or yield constraints indifferently; these are in fact the same type of constraints, but set differently.

[0122] By "yield of a collected fraction ", within the meaning of the present invention, means the yield of a target species contained in said collected fraction.

[0123] In certain embodiments, the method according to the invention also comprises a step of modifying the volume of mixture to be separated injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies).

[0124] For example, in the case of a binary blend, if both the extract and raffinate purities are higher than the predetermined purities, this means that the purities are beyond the specifications; the blend quantity can be increased to " degrade »purities up to specifications. If both the extract and raffinate purities are below the predetermined purities, this means that the purities are below specifications; the amount of mixing can be decreased to improve separation and increase the purities up to specifications. This mode of operation can be adjusted when only one purity is of interest. As noted above, one can also measure a quantity of the target species and its yield related to the presence of target species lost in other collections. This yield is directly related to the purity of the other fractions.

[0125] Preferably, the steps of measuring the purity (and / or yield) of at least one collected fraction, of comparing the measured purity(ies) (and / or the measured yield(s)) with a target purity (and / or a target yield), and of modifying the volume of mixture to be separated injected per cycle are carried out at least partly in parallel with the steps of detecting the history of the low concentration characteristic point, comparing its position with a target position and adjusting the carrying volume of the characteristic point.

[0126] Changing the amount of mixture to be separated can be implemented by changing the volume of mixture injected over a period. Changing the volume of mixture to be separated can be done in several ways: by increasing or reducing the mixture injection rate; and / or by increasing or reducing the period duration.

[0127] Controlling the quantity of mixture to be separated (or charge) to be injected requires measuring at least one purity and / or yield data item. The following non-limiting example illustrates a method for defining the usable quantity of charge to be injected if two purities are targeted for the extract and the raffinate. Consider the following variables: P* ext: Target purity of the extract; P* raff: Target purity of the raffinate; P ext: Measured purity of the extract; P raff: Measured purity of the raffinate; ε ext = P ext - P* ext , i.e. the difference between the measured purity and the target purity of the extract ε raff = P raff - P* raff , i.e. the difference between the measured purity and the target purity of the raffinate.

[0128] Purities can be calculated by determining the mass or molar concentrations of one or more species of interest in the fraction concerned.

[0129] According to a first method, PID regulations can be carried out on a quantity obtained from the deviations of a single purity or purities from their target value (ε ext and / or ε raff ). Another possibility is to use the value of a function dependent on the purities obtained and the target purities to calculate the new quantity to be injected.

[0130] Changing the injected quantity of mixture to be separated causes a disturbance that can cause the low concentration characteristic point to move away from the target position. The position of the low concentration characteristic point is then changed by adjusting the carrier volume of the characteristic point as already described above.

[0131] In the present invention, the regulation of the position of the low concentration characteristic point is carried out by controlling a single carrier volume chosen from the zone 1 volume, the zone 4 volume or a combination of the two (for example the average of the characteristic points of each zone). The distribution of the carrier volume between zone 1 and zone 4 remains precisely controlled, which makes it possible to respect high and / or low threshold values of the mobile phase volume. If the high and low thresholds are equal, the mobile phase volume injected per cycle is regulated according to the invention to a precise set value. This set value is naturally between the maximum and minimum limits of the mobile phase volume.The control method according to the invention therefore makes it possible to impose non-crossings of high and / or low thresholds and also to precisely control the volume of mobile phase according to criteria other than compliance with the front positions in zones 1 and 4.

[0132] In a more general description of the invention, it is possible to control the volume of mobile phase between maximum and minimum limits which are the extremes never to be exceeded by using intermediate control variables which are high and low thresholds. Two extreme ways of use are possible: • the upper and lower thresholds are set as being close to the maximum and minimum limits and the mobile phase volume is controlled according to the invention when its value goes outside the authorized range; • the upper and lower thresholds are modified according to the purity and / or yield measurements, the values of the thresholds themselves remaining constantly between the maximum and minimum limits of the mobile phase volume.

[0133] Thus, in certain embodiments, the method according to the invention comprises a step of modifying the setpoint of the volume of mobile phase injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0134] For example, in the case of a binary mixture, if both the extract and raffinate purities are higher than the predetermined purities, this means that the purities are beyond the specifications; the mobile phase volume can be decreased. If both the extract and raffinate purities are lower than the predetermined purities, this means that the purities are below the specifications; the mobile phase volume can be increased. This mode of operation can be adjusted when only one purity is of interest. Alternatively, a quantity of the target product can be measured and its yield related to the presence of target product lost in the other collections. This yield is directly related to the purity of the other fractions.

[0135] Preferably, the steps of measuring the purity (and / or yield) of at least one collected fraction, of comparing the measured purity(ies) (and / or the measured yield(s)) with a target purity (and / or with a target yield), and of modifying the volume of mobile phase injected per cycle are carried out at least partly in parallel with the steps of detecting on the history of the basic concentration characteristic point, of comparing its position with a target position and of adjusting the carrier volume of the characteristic point.

[0136] In some embodiments, during the step of modifying the volume of mobile phase injected per cycle, the volume of mixture to be separated injected per cycle remains constant.

[0137] In other embodiments, the volume of mixture injected per cycle and the volume of mobile phase injected per cycle are jointly varied based on the difference between the measured purities and the target purities (and / or the difference between the measured yield(s) and the target yield(s).

[0138] Any variation in the volume of mobile phase injected is carried out taking into account the maximum limit and / or the minimum limit for this volume which is or are provided in the method of the invention.

[0139] In some embodiments, the method comprises a step of defining the target position of the low concentration characteristic point as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s). This step makes it possible to improve the method already described, so as to optimize the target position with respect to the desired purities. The measured purities may be the purity of the extract and / or the raffinate.

[0140] Preferably, the steps of measuring the purity (and / or yield) of at least one collected fraction, of comparing the measured purity(ies) (and / or the measured yield(s)) with a target purity (and / or with a target yield), and of defining the target position are carried out at least partly in parallel with the steps of detecting on the history of the low concentration characteristic point, of comparing its position with a target position and of adjusting the carrier volume of the characteristic point.

[0141] The target position of the low concentration feature point can be such that it is sufficiently distant (in time) from the raffinate and extract collections, if the purity of both fractions (in case of binary mixture) is of importance. For example, the low concentration feature point can be positioned in the eluent injection step. However, it may be that the purity of only one fraction is of importance. In this case the target position can be close to either the raffinate or extract collections. For example, the low concentration feature point can be positioned in zone 1 or in zone 4.

[0142] As indicated above, one can also measure a quantity of the target product and its yield related to the presence of target product lost in other collections. This yield is directly related to the purity of the other fractions.

[0143] It should be noted that the measurement of the purity of the fractions can be obtained with a certain delay. However, this is not penalizing for the process because the steps of measuring the purity of at least one collected fraction, comparing the measured purity(ies) with a target purity, and defining the target position are advantageously carried out in parallel with the steps of detecting on the history of the basic concentration characteristic point, comparing its position with a target position and adjusting the carrier volume of the characteristic point. The definition of the new target position can then be implemented at each new available measurement or even every cycle by simply considering the last available measurements.

[0144] During the analysis time of the purity(ies), the process continues to adjust the carrier volume of the low concentration characteristic point by comparing the position of the characteristic point with the currently valid target position.

[0145] In order to define the target position of the characteristic point, several methods are possible.

[0146] According to a first method, a PID type regulation is applied to the parameters ε ext or ε raff . Thus if ε ext is negative, then the target position will be far from the extract line (the target position is closer to the raffinate line). Similarly, if ε ext is positive, then the target position will be brought closer to the extract line (the target position is moving away from the raffinate line). The regulation is similar if applied to ε raff . If ε raff is negative, then the target position will be far from the raffinate line (the target position is closer to the extract line). Similarly, if ε raff is positive, then the target position will be brought closer to the raffinate line (the target position is moving away from the extract line).

[0147] According to a second method, a PID regulator is applied to the combination of the deviations ε ext and ε raff and more generally to the combination of the deviations between the measured purities and / or yields and the target purities and / or yields.

[0148] According to a third method, a function dependent on the obtained purities and the target purities is directly used to calculate the new position of the characteristic point.

[0149] During the target position definition step, the target position can also be defined relative to the mobile phase injection position on the history. This allows adaptation to changes in the injection point position.

[0150] The optimization of the target position of the low concentration characteristic point can be carried out automatically.

[0151] In certain advantageous embodiments, the method further comprises: - the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; - the detection on the history of another characteristic point, called the high concentration characteristic point in the present description, located between the start of a raffinate collection step and the end of the following extract collection step; - the comparison of the position of the high concentration characteristic point with respect to a target position; - the adjustment of the carrier volume of the high concentration characteristic point modifying its position to bring it closer to its target position.

[0152] The concentration representative variable, observation node, and history may each independently be the same variable, observation node, and history as used for low concentration feature point detection; or may be a different variable, observation node, and history.

[0153] This characteristic point of high concentration is located between the raffinate and extract lines, i.e. in zone 2 or in zone 3, or at the interface of zones 2 and 3 (i.e. at the average injection position of the mixture to be separated), where the concentrations are high.

[0154] Document WO 2007 / 101944 describes in detail the detection and use of this characteristic high concentration point, which is represented in particular in the figures 6 to 11 And 18 à 21 from document WO 2007 / 101944.

[0155] What has been stated above about the characteristic point of low concentration applies by analogy to the characteristic point of high concentration.

[0156] The carrier volume that is adjusted to control the position of the high concentration characteristic point is for example the zone volume 2 and any variation in the zone volume 2 is reflected identically on the zone volume 3. Mobile phase volume constraint

[0157] In the method according to the first aspect, corresponding to the invention, the volume of the mobile phase injected per cycle is kept greater than or equal to a minimum limit and / or less than or equal to a maximum limit, at least during part of the method; and preferably during the entire method.

[0158] In the case where a maximum limit is imposed, when the volume of mobile phase injected per cycle reaches this maximum limit, it can no longer increase. Advantageously, when the volume of mobile phase reaches the maximum limit, it remains constant at this maximum limit.

[0159] In the case where a minimum limit is imposed, when the volume of mobile phase injected per cycle reaches this minimum limit, it can no longer decrease. Advantageously, when the volume of mobile phase reaches the minimum limit, it remains constant at this minimum limit.

[0160] In some embodiments, the mobile phase volume is maintained both greater than or equal to a minimum bound and less than or equal to a maximum bound. In some embodiments, the maximum bound and the minimum bound are the same. In this case, the volume of injected mobile phase is controlled at a constant volume equal to the minimum (or maximum) bound. Triggering of the regulation according to the invention during the process

[0161] Process control by reference to the low concentration characteristic point may in some cases be implemented during the process, when a condition is reached, and not necessarily from the start of the process.

[0162] For example, this regulation can be triggered when the volume of injected mobile phase reaches or exceeds a certain threshold.

[0163] Thus, another object of the invention is a method for separating a mixture in a system comprising a set of one or more chromatography columns, the method successively comprising, in a cyclic manner: a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract and a step of injecting mobile phase; the volume of the mobile phase injected per cycle being maintained greater than or equal to a minimum limit and / or less than or equal to a maximum limit; the method being such that: as long as the volume of the mobile phase injected per cycle is greater than or equal to a threshold value or less than or equal to a threshold value, the method comprises: ▪ the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; ▪ the detection on the history of a first characteristic point between the start of the extract collection step and the end of the following mobile phase injection step (also called the desorption characteristic point); ▪ the detection on the history of a second characteristic point between the start of the mobile phase injection step and the end of the following raffinate collection step (also called the adsorption characteristic point);▪ comparing the position of each of the characteristic points to a respective target position; ▪ adjusting the carrier volume of the first characteristic point and the carrier volume of the second characteristic point, modifying the position of the first characteristic point and the second characteristic point, to bring them closer to their respective target positions; when the volume of the mobile phase injected per cycle reaches a threshold value, the method is a method as described above: that is to say, the regulation based on the first characteristic point and on the second characteristic point is abandoned, in favor of the regulation based on the low concentration characteristic point described above. ;

[0164] For example, the threshold value may be the minimum and / or maximum bound of the injected mobile phase. Alternatively, it may be a threshold value having a predetermined deviation from the minimum and / or maximum bound.

[0165] Thus, as long as the mobile phase volume injected per cycle has not reached the threshold value, the chromatographic process is controlled by comparing the position of two adsorption and desorption characteristic points with a respective target position. This allows for optimal control accuracy. When the mobile phase volume, changed due to control or for any other reason, reaches the threshold value (and for example minimum limit or maximum limit), the process is then controlled less precisely using the low concentration characteristic point, as described above, which avoids the disadvantages associated with too great an increase or too great a decrease in the injected mobile phase.

[0166] The regulation by means of the two characteristic points of adsorption and desorption has been described in detail in WO 2007 / 101944 as mentioned above.

[0167] Furthermore, what has been stated above about the low concentration characteristic point applies by analogy to each of the adsorption and desorption characteristic points. Computer program

[0168] Another subject of the invention consists of a computer program comprising program code instructions for executing the steps of the method according to the invention when said program is executed on a computer.

[0169] The invention also relates to a computer-readable storage medium on which a computer program as defined above is recorded.

[0170] The invention also relates to a system comprising a processor coupled to a memory on which is recorded a computer program as defined above. Said system can also comprise the chromatographic separation system as described above or be only a control system, connected to the separation system and separate from it. Measurement of purities and / or yields by online detectors

[0171] The present disclosure also describes, according to a second aspect not representing the invention, a method for separating a mixture in a system comprising a plurality of chromatography columns, the method successively comprising, in a cyclic manner, in a given part of the system: a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract and a step of injecting mobile phase; the method further comprising measuring the purity and / or yield of at least one collected fraction chosen from the extract and the raffinate, said purity and / or yield measurement comprising the following steps: determining, at a node of the system, histories of at least two respective variables representative of the concentration of at least two species contained in the mixture to be separated, by means of at least one rapid online detector; determining the concentration of at least two species of the mixture to be separated in the fraction collected from the histories; determining the purity and / or yield of the fraction collected from the concentrations.

[0172] By “determination of histories of at least two respective variables representative of the concentration of at least two species contained in the mixture to be separated", we mean the determination of histories of at least two variables, these variables each being representative of the concentration of at least two species (i.e. of the set of at least two species) contained in the mixture to be separated.

[0173] Purity corresponds to the ratio of the concentration of one or more species in the collected fraction, compared to the sum of all the concentrations determined in the collected fraction.

[0174] The yield corresponds to the ratio of the quantity of a species in one of the two collected fractions compared to the total quantity of this species in the two collected fractions combined.

[0175] By " online detector”, for the purposes of the present disclosure, a detector positioned at the outlet of a chromatographic column, i.e. on a connection line between two successive columns is understood. By "online detector",we also mean a bypass detector whose sample is positioned at the outlet of a chromatographic column, i.e. on a connection line between two successive columns. Thus, an in-line detector is not located on an outlet line of the chromatographic system (i.e. on a collection line for the extract or raffinate).

[0176] According to the method according to the second aspect, a single or, alternatively, several detectors may be used. In the case where at least two detectors are used, they may be positioned at the outlet of the same column, or at the outlet of different columns.

[0177] In a system in which a cyclic chromatographic process is implemented, two types of detectors can be used: fast detectors and slow detectors. In this method according to the second aspect, the at least one online detector used is a fast detector.

[0178] By "we mean fast detector " within the meaning of this disclosure a detector whose response time is less than one twentieth of the duration of a cycle. By "slow detector" a detector whose response time is greater than one twentieth of the duration of a cycle.

[0179] A fast detector emits a signal dependent on the concentration profile of the compounds moving inside the detector. It can therefore make it possible to detect a characteristic point on a history, for example for the implementation of the separation method according to the first aspect, corresponding to the invention.

[0180] Examples of rapid detectors include UV / visible absorbance detectors, measuring at single or multiple wavelengths, colorimeters, density meters, conductivity meters, refractometers, brixmeters, polarimeters, nuclear magnetic resonance devices, and spectrometers. NIR (near infrared), IR(infrared), FTIR (Fourier transform infrared) and Raman. A temperature detector can be used to correct the signal given by the detectors mentioned above.

[0181] In certain embodiments, the at least one rapid online detector is chosen from the rapid detectors mentioned above, provided that the at least two variables representative of the concentration of at least two species contained in the mixture to be separated detected are different.

[0182] The at least one detector can measure one or more variables representative of the concentration of at least two species contained in the mixture to be separated. Thus, for example, spectrometric type detectors, after calibration of the absorption or emission wavelengths to the species to be separated, will be able to measure the concentration of said species with a single device.

[0183] The use of a single rapid detector sometimes does not allow for the precise measurement of a species concentration.

[0184] As illustrated in Example 6, the use of a combination of a polarimeter and a densimeter during the purification of a mixture comprising glucose, fructose and glucose polymers in minor quantities makes it possible to obtain variables representative of the concentrations of the species in the mixture: the densimeter gives information on the sum of all the concentrations and the polarimeter measures a rotation of the polarized light, knowing that glucose and glucose polymers have a positive contribution while fructose has a negative contribution. In this case, the representative variables are density and polarity. The combination of the values of these variables obtained by the two detectors makes it possible to evaluate the concentrations of species in the mixture (after calibration of the detectors).

[0185] In the case of using a spectrometer (IR, NIR, FTIR, Raman, etc.), each measurement of the spectrometer returns an information vector consisting of a set of absorption or emission values at different wavelengths. The adsorption or emission at these different wavelengths is each potentially a variable representing a concentration of the species. Indeed, a combination of these adsorption or emission values together makes it possible to estimate the value of the concentrations of one or more of the species. The determination of such a combination is done by standard chemometric tools. In embodiments, at least two fast online detectors are used to determine the histories. Preferably, they are chosen from the fast detectors mentioned above.

[0186] In some embodiments, the at least two in-line detectors are a polarimeter and a density meter.

[0187] According to other embodiments, the at least two in-line detectors are a density meter and a conductivity meter.

[0188] In some embodiments, the at least two species of the mixture to be separated are monosaccharides. In these embodiments, the extract and the raffinate are enriched in different monosaccharides. In particular, the monosaccharides are selected from the monosaccharides mentioned above.

[0189] In some embodiments, the at least two species of the mixture to be separated are glucose and fructose. In these embodiments, the mixture to be separated may contain only glucose and fructose, optionally diluted in a solvent, for example water. Alternatively, the mixture to be separated may contain one or more other compounds, such as glucose polymers.

[0190] Advantageously, when the at least two species of the mixture to be separated are glucose and fructose, at least two rapid online detectors are used, preferably a polarimeter and a densimeter.

[0191] According to other embodiments, the at least two species of the mixture to be separated are an ionized species, for example in salt form, and a non-ionized species. In these embodiments, the extract and the raffinate are enriched in different species, that is to say that the extract is enriched in ionized species and the raffinate is enriched in non-ionized species or vice versa (the extract is enriched in non-ionized species and the raffinate is enriched in ionized species).

[0192] Examples of ionized species include amino acids, salts, or organic acids found in ionized form at the pH and operating temperature of the chromatographic separation.

[0193] As non-ionized species at the pH and operating temperature of the chromatographic separation, alcohols and sugars, such as the monosaccharides and polysaccharides mentioned above, can be mentioned.

[0194] Advantageously, when the at least two species of the mixture to be separated are an ionized species and a non-ionized species, at least two rapid in-line detectors are used, preferably a densimeter and a conductivity meter.

[0195] In certain embodiments, the method comprises, after the step of determining histories of at least two respective variables representative of the concentration of at least two species contained in the mixture to be separated, a step of determining the average value of each variable over a measurement interval of the histories.

[0196] According to other embodiments, the method comprises, after the step of determining histories of at least two respective variables representative of the concentration of at least two species contained in the mixture to be separated, a step of determining the value of the integration of each variable over a measurement interval of the histories.

[0197] In the above, each historical measurement interval can be defined in its entirety by the collection of the fraction in question. In other words, said historical measurement interval corresponds, during a cycle, to the times (or volumes) at which the collection line of the fraction in question is located between the same successive columns as the detector in question.

[0198] The said measurement interval may also correspond to only part of the collection of the fraction in question.

[0199] Said measurement interval may also correspond to part or all of the collection of the fraction in question, and to a portion of the history immediately following the collection of the fraction in question.

[0200] Said measurement interval may also correspond to part or all of the collection of the fraction in question, and to a portion of the history immediately preceding the collection of the fraction in question.

[0201] These last three embodiments are particularly useful if an asymmetry is present in the chromatography system.

[0202] This measurement of purities and / or yields can be used with all state-of-the-art control processes requiring a purity measurement to carry out actions.

[0203] In some embodiments, the method further comprises: ▪ the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; ▪ the detection on the history of a characteristic point of high concentration located between the start of a raffinate collection step and the end of the following extract collection step; ▪ the comparison of the position of the characteristic point of high concentration with respect to a target position; ▪ the adjustment of the carrier volume of the characteristic point of high concentration modifying the position of the characteristic point of high concentration to bring it closer to its target position.

[0204] What has been stated above about the characteristic point of high concentration can thus be applied to the second aspect described here.

[0205] In some embodiments, the method further comprises: ▪ detecting on the history of a first characteristic point between the start of the extract collection step and the end of the following mobile phase injection step (also called desorption characteristic point); ▪ detecting on the history of a second characteristic point between the start of the mobile phase injection step and the end of the following raffinate collection step (also called adsorption characteristic point); ▪ comparing the position of each of the characteristic points to a respective target position; ▪ adjusting the carrier volume of the first characteristic point and the carrier volume of the second characteristic point, modifying the position of the first characteristic point and the second characteristic point, to bring them closer to their respective target positions.

[0206] What has been stated above about the characteristic points of adsorption and desorption can thus be applied to the second aspect described here.

[0207] In some embodiments, the method further comprises: ▪ determining, at a node of the system, the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; ▪ detecting on said history a characteristic point of low concentration between the start of a step of collecting the extract and the end of the following step of collecting the raffinate; ▪ comparing the position of the characteristic point of low concentration with a target position; ▪ adjusting the carrier volume of the characteristic point of low concentration, modifying the position of the characteristic point of low concentration to bring the position of the characteristic point of low concentration closer to its target position; ▪ preferably, the volume of the mobile phase injected per cycle being kept greater than or equal to a minimum limit and / or less than or equal to a maximum limit.

[0208] What has been stated above about the characteristic point of low concentration can thus be applied to the second aspect described here.

[0209] In the above embodiments, the histories on which the high concentration characteristic point, the adsorption and desorption characteristic points, the low concentration characteristic point are detected, and the histories from which the concentration of at least two species of the mixture to be separated in at least one collected fraction is determined may independently be the same histories or different histories. Il the same applies to the variables representing the concentration of species contained in the mixture to be separated.

[0210] In some embodiments, the method also comprises a step of comparing the measured purity and / or the measured yield with a target purity and / or a target yield. If the purity and / or yield are measured in the extract and in the raffinate, the method may comprise a step of comparing the two measured purities and / or yields with their respective target purities and / or yields.

[0211] In certain embodiments, the method also comprises a step of modifying the volume of mixture to be separated injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0212] In some embodiments, the method comprises a step of modifying the volume of mobile phase injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0213] In some embodiments, the volume of mixture injected per cycle and the volume of mobile phase injected per cycle are jointly varied based on the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0214] In some embodiments, the method comprises a step of defining the target position of the characteristic point(s) as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s). This may involve defining the target position of the high concentration characteristic point and / or defining the target position of the adsorption characteristic point and / or that of the desorption characteristic point and / or that of the low concentration characteristic point.

[0215] What has been stated above about comparing a measured purity and / or yield with a target purity and / or yield, modifying, as a function of this comparison, the injected mixture volume and the mobile phase volume and defining, as a function of this comparison, the target position of the characteristic point can thus be applied to the second aspect described here.

[0216] In some embodiments of the method, quantities such as the average operating velocity of the fluid in the columns may also be modulated if it is desired to modify the production of the system or to control the pressure on one or more columns of the system.

[0217] Another subject matter described in the present disclosure (not representing the invention) consists of a computer program comprising program code instructions for carrying out the steps of the method according to the second aspect when said program is executed on a computer.

[0218] Also described in the present disclosure (without representing the invention) is a computer-readable storage medium on which is recorded a computer program as defined above.

[0219] Also described in the present disclosure (without representing the invention) is a system comprising a processor coupled to a memory on which is recorded a computer program as defined above. Said system may also comprise the chromatographic separation system as described above or be only a control system, connected to the separation system and separate from it. Measurement of purities and / or yields on an intermediate tank

[0220] The present disclosure also describes, according to a third aspect not representing the invention, a method of separating a mixture in an installation comprising: a first system comprising a plurality of chromatography columns, an outlet line for collecting a raffinate and an outlet line for collecting an extract; at least one second system placed downstream of the first system; and at least one tank fed by one of said outlet lines of the first system, and feeding the second system, preferably with a continuous flow; the method comprising successively, cyclically, in a given part of the first system: a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract and a step of injecting mobile phase; the method further comprising the measurement of the purity and / or yield of at least one collected fraction chosen from the extract and the raffinate, said measurement of purity and / or yield comprising the following steps: determining the concentration of at least two species of the mixture to be separated in the fraction collected in the tank; determining the purity and / or yield of at least one species of the fraction collected in the tank.

[0221] The determination of the purity and / or yield of at least one species of the fraction collected in the tank is carried out from the concentrations of the at least two species of the mixture to be separated determined in the fraction collected in the tank.

[0222] Purity corresponds to the ratio of the concentration of one or more species in the fraction collected in the tank, compared to the sum of all the concentrations determined in the fraction collected in the tank.

[0223] The yield corresponds to the ratio of the quantity of a species in one of the two collected fractions compared to the total quantity of this species in the two collected fractions combined.

[0224] The at least one second system preferably operates with a continuous feed and at a stable flow rate and composition. The tank ensures a continuous feed to the second system in the event of a short shutdown of the first system. The presence of the tank can, for example, allow one hour of operation of the second system in the event of a shutdown of the chromatography installation. The tank also makes it possible to standardize the flow rate and smooth the composition of the collected fraction before its feed to the second system in the event of asymmetry in the first system from one column to the other. Preferably, the tank is periodically fed with a collected fraction and constantly emptied to the second system. It is never purged or even completely rinsed between each cycle of the system.

[0225] In the case of an equilibrium chromatography system, the fraction contained in the tank has an average composition very close to the average composition of the fraction collected over a cycle.

[0226] When the chromatography system is modified or evolving, the composition of the fraction contained in the tank is not strictly equal to the composition of the fraction directly collected over a cycle of the system, the composition of the fraction contained in the tank evolving with a delay compared to the composition of the directly collected fraction due to the duration of the renewal of the tank. The renewal of the contents of the tank is all the more delayed as the volume of the tank is large. This measurement remains, however, sufficiently representative, in particular to allow regulation of the chromatography system.

[0227] In this description, the terms " output line " And " collection line » have the same meaning.

[0228] In some embodiments, the determination of the concentration of at least two species of the mixture to be separated in the fraction collected in the tank is carried out using at least one detector, preferably a slow detector.

[0229] Examples of slow detectors include near-infrared spectrometers, infrared spectrometers, Fourier transform infrared spectrometers, and Raman spectrometers. In cases where the acquisition of several spectra is necessary to obtain a signal for accurate measurement, the total acquisition time can be significant. Examples of slow detectors include all nuclear magnetic resonance techniques.

[0230] According to other embodiments, the determination of the concentration of at least two species of the mixture to be separated in the fraction collected in the tank is carried out using an analytical chromatography system, such as high performance liquid chromatography (HPLC), high pressure liquid chromatography (UPLC), ultra high pressure liquid chromatography (UHPLC), gas chromatography (GC).

[0231] The second system may in particular be a chromatography unit, an enzymatic transformation unit, a chemical transformation unit, a distillation unit, a membrane concentration unit, or an evaporation unit. In certain embodiments, the second system is an evaporation unit.

[0232] The installation includes a tank (or "intermediate tank") fed by an outlet line. The tank can therefore be fed by the raffinate outlet line, or by the extract outlet line; respective tanks can be fed by each outlet line (i.e. the raffinate outlet line and the extract outlet line). The tank is fed by the fraction collected in the outlet line on which it is present. The tank is connected, directly or indirectly, to a second system which is fed by the fraction contained in the tank.

[0233] Preferably, the tank has a volume less than the volume of the collected fraction (i.e., the collected fraction feeding it) over a cycle. In some embodiments, the tank contains a fraction volume equal to the volume of the collected fraction over a period of one to five periods, preferably three periods.

[0234] Particularly advantageously, when several purity and / or yield measurements are carried out successively, the tank is not cleaned and / or purged and / or rinsed between these measurements.

[0235] The mixture to be separated may contain the species already listed above.

[0236] In some embodiments, the at least two species of the mixture to be separated whose concentration is determined are monosaccharides. In these embodiments, the extract and the raffinate are enriched in different monosaccharides. In particular, the monosaccharides are chosen from the monosaccharides mentioned above in the description of the first aspect corresponding to the invention.

[0237] In some embodiments, the at least two species of the mixture to be separated are glucose and fructose.

[0238] In some embodiments, the method further comprises: ▪ the determination, at a node of the system, of the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; ▪ the detection on the history of a characteristic point of high concentration located between the start of a raffinate collection step and the end of the following extract collection step; ▪ the comparison of the position of the characteristic point of high concentration with respect to a target position; ▪ the adjustment of the carrier volume of the characteristic point of high concentration modifying the position of the characteristic point of high concentration to bring it closer to its target position.

[0239] What has been stated above about the characteristic point of high concentration can be applied to the third aspect described here.

[0240] In some embodiments, the method further comprises: ▪ detecting on the history of a first characteristic point between the start of the extract collection step and the end of the following mobile phase injection step (also called desorption characteristic point); ▪ detecting on the history of a second characteristic point between the start of the mobile phase injection step and the end of the following raffinate collection step (also called adsorption characteristic point); ▪ comparing the position of each of the characteristic points to a respective target position; ▪ adjusting the carrier volume of the first characteristic point and the carrier volume of the second characteristic point, modifying the position of the first characteristic point and the second characteristic point, to bring them closer to their respective target positions.

[0241] What has been stated above about the characteristic points of adsorption and desorption can be applied to the third aspect described here.

[0242] In some embodiments, the method further comprises: ▪ determining, at a node of the system, the history of a variable representative of the concentration of one or more species contained in the mixture to be separated; ▪ detecting on said history a characteristic point of low concentration between the start of a step of collecting the extract and the end of the following step of collecting the raffinate; ▪ comparing the position of the characteristic point of low concentration with a target position; ▪ adjusting the carrier volume of the characteristic point of low concentration, modifying the position of the characteristic point of low concentration to bring the position of the characteristic point of low concentration closer to its target position; ▪ preferably, the volume of the mobile phase injected per cycle being kept greater than or equal to a minimum limit and / or less than or equal to a maximum limit.

[0243] What has been stated above about the characteristic point of low concentration can be applied to the third aspect described here.

[0244] In some embodiments, the method also comprises a step of comparing the measured purity and / or the measured yield with a target purity. If the purity and / or the yield are measured in the extract and in the raffinate, the method may comprise a step of comparing the two measured purities and / or yields with a respective target purity and / or yield.

[0245] In certain embodiments, the method also comprises a step of modifying the volume of mixture to be separated injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0246] In some embodiments, the method comprises a step of modifying the volume of mobile phase injected per cycle as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0247] In some embodiments, the volume of mixture injected per cycle and the volume of mobile phase injected per cycle are jointly varied based on the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s).

[0248] In some embodiments, the method comprises a step of defining the target position of the characteristic point(s) as a function of the difference between the measured purity(ies) and the target purity(ies) (and / or the difference between the measured yield(s) and the target yield(s). This may involve defining the target position of the high concentration characteristic point and / or defining the target position of the adsorption characteristic point and / or that of the desorption characteristic point and / or that of the low concentration characteristic point.

[0249] What has been stated above about comparing a measured purity and / or yield with a target purity and / or yield, changing, as a function of this comparison, the injected mixture volume and the mobile phase volume and defining, as a function of this comparison, the target position of the characteristic point can be applied to the third aspect described here.

[0250] Another subject matter described in the present disclosure (not representing the invention) consists of a computer program comprising program code instructions for carrying out the steps of the method according to the third aspect when said program is executed on a computer.

[0251] Also described in the present disclosure (without representing the invention) is a computer-readable storage medium on which is recorded a computer program as defined above.

[0252] Also described in the present disclosure (without representing the invention) is a system comprising a processor coupled to a memory on which is recorded a computer program as defined above. Said system may also comprise the chromatographic separation system as described above or be only a control system, connected to the separation system and separate from it. EXAMPLES

[0253] The following examples illustrate the aspects described in the present disclosure without limiting them. Examples 1 to 5 and 7 (according to the invention) illustrate the first aspect and Example 6 (not according to the invention) illustrates the second aspect.

[0254] In Examples 1 to 5 below, a chromatographic simulation model was used with the following parameters: species present in the mixture to be separated: glucose polymers of order greater than or equal to 2, glucose and fructose; stationary phase: XA2004 / 30 Ca resin from Novasep; mobile phase: water; elution temperature: between 55°C and 65°C.

[0255] Concentration graphs representing a loading pulse, high concentration saturation and low concentration saturation, in a 2 meter long and 8.5 cm diameter column, at a flow rate of 20 mL / min, are shown respectively in figures 5 .A, 5.B and 5.C. These data allow the reconstruction of the model used, using any chromatographic simulation software, for example using the methods described in the manual Preparative Chromatography of Fine Chemicals and Pharmaceutical Agents, Henner Schmidt-Traub, Wiley-VCH, ISBN-13 978-3-527-30643-5.

[0256] The method applied in examples 1, 2, 4 and 5 is an SSMB4 with a total column height of 8 meters.

[0257] The method applied in Example 3 is an SMB with a total column height of 8 meters.

[0258] In Examples 6 to 7 below, an actual pilot chromatographic separation was performed, with the following parameters: species present in the mixture to be separated: glucose polymers of order greater than or equal to 2, glucose and fructose; stationary phase: XA2004 / 30 Ca resin from Novasep; mobile phase: water; elution temperature: between 55°C and 65°C; the pilot is composed of 4 columns 2 meters long and 8.5 cm in diameter; the flow rates of the products and eluent are between 20 and 30 mL / min; the process applied is an SSMB with a total column height of 8 meters. Example 1

[0259] A chromatographic process simulation as described above is carried out.

[0260] Between cycle 1 and cycle 10, the system is stabilized, the volumes of the different zones are fixed and kept constant. The volume of mobile phase injected is 0.18 BV (bed volume).

[0261] From cycle 10, the zones are regulated with target positions of characteristic points located at + 0.17 for zone 4 (deviation of the target position of the adsorption characteristic point in volume related to the cycle volume, compared to the average eluent injection position) and -0.17 for zone 1 (deviation of the target position of the desorption characteristic point in volume related to the cycle volume, compared to the average eluent injection position). This corresponds to a target position of the low concentration characteristic point located on the average eluent injection point.

[0262] A comparison is then made between a process regulated using the two characteristic points of adsorption and desorption, without limitation of mobile phase volume (as described in document WO 2007 / 101944), and a process regulated using the low concentration characteristic point according to the invention in which a maximum volume threshold has been imposed.

[0263] The results are summarized in the figures 6 , 7 , 8 And 9 .

[0264] It is found that when the process is controlled using both adsorption and desorption characteristic points, both characteristic points each reach their target position. The same is true for the low concentration characteristic point. In order to achieve these characteristic point position targets, the mobile phase volume is increased to 0.27 BV.

[0265] In the second case, the mobile phase volume is limited to 0.22 BV and the chromatographic process is controlled according to the invention using the low concentration characteristic point. The low concentration characteristic point is unchanged. It can be seen that the target position of the low concentration point is reached, the characteristic point reaches the position of the eluent injection point. However, the target positions of the adsorption and desorption characteristic points are not reached. The mobile phase volume required to reach these target positions is not reached and has been limited to the predefined maximum value. The adsorption and desorption characteristic points are located at + 0.143 and - 0.143 relative to the average eluent injection position (in volume relative to the cycle volume) respectively.The absolute values of the deviations between each of the characteristic adsorption and desorption points and their target position are identical and the sum of the deviations is zero.

[0266] Thus, the method according to the invention makes it possible not to exceed a high limit of the volume of mobile phase used, which is not possible with a regulated method taking into account two characteristic points of adsorption and desorption. Example 2

[0267] A chromatographic process simulation as described above is carried out.

[0268] Between cycle 1 and cycle 10, the system is stabilized, the volumes of the different zones are fixed and kept constant. The mobile phase volume is 0.25 BV.

[0269] From cycle 10, the zones are regulated with target positions of characteristic points located at + 0.10 for zone 4 (position of the characteristic adsorption point in volume related to the cycle volume, compared to the average eluent injection position) and - 0.10 for zone 1 (position of the characteristic desorption point in volume related to the cycle volume, compared to the average eluent injection position). This corresponds to a target position of the low concentration characteristic point located on the average eluent injection point.

[0270] A comparison is then made between a process controlled using the two characteristic points of adsorption and desorption, without limitation of mobile phase volume (as described in document WO 2007 / 101944), and a process controlled using the low concentration characteristic point according to the invention in which a minimum volume threshold has been imposed.

[0271] The results are summarized in the figures 10 , 11 , 12 And 13 .

[0272] It is observed that when the process is controlled using the two adsorption and desorption characteristic points, the two characteristic points each reach their target position (at + 0.10 and - 0.10). The same is true for the low concentration characteristic point. In order to achieve these characteristic point position targets, the mobile phase volume is reduced to 0.18 BV.

[0273] In the second case, the mobile phase volume is minimized to 0.20 and the chromatographic process is controlled according to the invention using the low concentration characteristic point. The low concentration characteristic point is unchanged. It can be seen that the target position of the low concentration point is reached, at the eluent injection point. However, the target positions of the adsorption and desorption characteristic points are not reached. The mobile phase volume to reach these target positions is not reached and has been limited to the predefined minimum value. The adsorption and desorption characteristic points are located at + 0.12 and - 0.12 relative to the average eluent injection position (in volume relative to the cycle volume). The deviations between each of the adsorption and desorption characteristic points and their target position are identical.

[0274] Thus, the method according to the invention makes it possible not to exceed a low limit of the volume of mobile phase used, which is not possible with a regulated method taking into account two characteristic points of adsorption and desorption. Example 3

[0275] A chromatographic process simulation as described above is carried out.

[0276] Measurements of purity and fructose yield in the extract and raffinate fraction are carried out while these fractions are collected in tanks placed at the chromatography outlet and with continuous withdrawal on each of them so that the average volume in the tanks represents three periods

[0277] Between cycle 1 and cycle 10, the system is stabilized, the volumes of the different zones are fixed and kept constant. The injected volume of mixture to be separated (charge) is 0.175 BV. The yield is 90.3% and the purity is 84.9%. The target position of the low concentration characteristic point is located at the average eluent injection point.

[0278] From cycle 10, the zones and the injected charge volume are regulated with purity targets of 90% and yield of 85% for the extract fraction. The mobile phase volume is kept constant at the value of + 0.2 BV using the regulation using the low concentration characteristic point of the process according to the invention.

[0279] The results are summarized in figure 14 .

[0280] The low concentration characteristic point is regulated at the average injection position of the eluent. The volume of the mobile phase is thus kept constant and controlled.

[0281] The yield and purity objectives are met and the injected charge volume is increased to +0.179 BV. Example 4

[0282] A chromatographic process simulation as described above is carried out.

[0283] Measurements of purity and fructose yield in the extract fraction are carried out.

[0284] Between cycle 1 and cycle 10, the system is stabilized, the volumes of the different zones are fixed and kept constant. The injected volume of mixture to be separated (charge) is 0.175 BV. The yield is 90.3% and the purity is 84.9%. The mobile phase volume is constant at the value of + 0.2 BV. The target position of the low concentration characteristic point is located at the average eluent injection point.

[0285] From cycle No. 10, the process is regulated using the low concentration characteristic point according to the process of the invention, with purity and yield objectives of 90% and 85% respectively and an injected charge volume kept constant at the value of + 0.175 BV.

[0286] The results are summarized in figure 15 .

[0287] The low concentration characteristic point is regulated at the mid-position of the eluent injection. The volume of the mobile phase is thus controlled.

[0288] The yield and purity targets are met and the mobile phase volume is reduced to +0.194 BV. Example 5

[0289] A chromatographic process simulation as described above is carried out.

[0290] Measurements of purity and fructose yield in the extract fraction are carried out.

[0291] The target position of the low concentration characteristic point is optimized based on the desired purity and yield.

[0292] In a first test, the target position of the low concentration characteristic point is set at - 0.1 BV (towards the extract) relative to the average injection position of the eluent and the regulation described in example 4 is applied.

[0293] The results are summarized in figure 16 . A mobile phase volume of + 0.198 BV is obtained.

[0294] In a second test, the target position of the low concentration characteristic point is set at + 0.1 BV (towards the raffinate) relative to the average eluent injection position and the regulation described in Example 4 is applied.

[0295] The results are summarized in figure 17 . A mobile phase volume of + 0.194 BV is obtained.

[0296] It is thus possible to position the characteristic point of low concentration to obtain the minimum mobile phase volume. With a characteristic point positioned at + 0.04 BV towards the raffinate relative to the average eluent injection position, the minimum mobile phase volume of this separation is obtained (at a value of + 0.193 BV) with an injected charge volume fixed at + 0.175 BV. The results are summarized in figure 18 . Example 6

[0297] An experimental chromatographic test as described above is carried out.

[0298] A chromatographic process is implemented to treat a mixture to be separated containing glucose, fructose and a small amount of glucose polymer (with a degree of polymerization of 2 (DP2), 3 (DP3) and above).

[0299] A calibration of a densimeter and a polarimeter is carried out for the two species to be separated (glucose and fructose), as a function of temperature and as a function of the concentration of the species. The results of the polarimeter calibration are summarized in Figure 19 and 20 .

[0300] Two histories are then measured using the previously calibrated densimeter and polarimeter positioned in line.

[0301] THE Figures 1 and 2illustrate the example because from these histories, over a cycle, an integration of the signal from each detector is carried out on the portion of the history corresponding to a fraction collection, for each fraction (the extract and the raffinate), in order to determine an “average” density and an “average” polarity for each fraction collected.

[0302] Glucose and the mixture of glucose polymers are dextrorotatory, that is, the rotation angle induced by glucose and glucose polymers measured by a polarimeter is positive. Fructose is levorotatory, that is, the rotation angle induced by fructose measured by a polarimeter is negative.

[0303] The rotation angle measured by a polarimeter during a measurement on a mixture is the sum of the rotation angles induced by the different species contained in the mixture in a unitary manner.

[0304] Thus, we can establish the following equation: α = α T Glu cos e . C Glu cos e + α T Fructose . C Fructose , in which: α is the angle of the glucose / fructose mixture measured by the polarimeter, α(T) is the rotation angle of the species at the temperature applied to the measurement, obtained during the calibration of the polarimeter, and C is the concentration of the species.

[0305] The measurement of the density of a mixture of glucose and fructose by a densimeter as a function of the concentrations of each sugar responds to the following equation: Densité = β T . C Glu cos e + C Fructose + Densité Eluant T , in which: Density is the density measured by the densimeter, Eluant Density (T) is the density of the eluent at the measurement temperature, β(T) is the density factor of glucose and fructose at the temperature applied to the measurement, this factor being identical for pure glucose and fructose at a given temperature, and C is the concentration of the species.

[0306] These two equations allow, from the average density and rotation angle characterizing each fraction, to calculate by the Gaussian pivot method the concentration of glucose and fructose in the mixture. The purity of each fraction in glucose and fructose can then be calculated from these concentrations.

[0307] This method allows the glucose and fructose concentrations of a collected fraction to be measured relatively accurately. The purities of the extract and raffinate can also be corrected to the extent of a correction term depending on the distribution of other minority impurities such as glucose polymers between these two collected fractions. Example 7

[0308] An experimental chromatographic test as described above is carried out.

[0309] Measurements of purity and fructose yield in the extract fraction are carried out.

[0310] Between cycle 1 and cycle 20, the system is stabilized and regulated. The injected volume of mixture to be separated (charge) is 0.15 BV. The yield is 90% and the purity is 90%. The target position of the low concentration characteristic point is located at the average eluent injection point.

[0311] From cycle 20, the zones and the injected charge volume are regulated with purity targets of 86% and yield of 92%. The mobile phase volume is kept constant at the value of + 0.18 BV using the regulation using the low concentration characteristic point of the process according to the invention. From cycle 60, oscillations on the concentration of the injected charge were carried out and the control process maintained the system performance.

[0312] The results are summarized in figure 21 .

[0313] The low concentration characteristic point is regulated at the average injection position of the eluent. The volume of the mobile phase is thus kept constant and controlled.

[0314] The yield and purity objectives are met and the injected charge volume is increased to +0.16 BV.

Claims

1. Method of separating a mixture in a system comprising a plurality of chromatography columns, the method successively comprising, in a cyclic manner, in a given part of the system: - a step of collecting a raffinate, a step of injecting the mixture to be separated, a step of collecting an extract, and a step of injecting a mobile phase; the system comprising areas 1, 2, 3 and 4, area 1 being located between a mobile phase injection line and a line for collecting the extract, area 2 being located between said line for collecting the extract and a line for injecting the mixture to be separated, area 3 being located between said line for injecting the mixture to be separated and a line for collecting the raffinate, and area 4 being located between said line for collecting the raffinate and said mobile phase injection line; wherein the method further comprises: - the determination, in a node of the system, of the history of a variable that is representative of the concentration of one or more species contained in the mixture to be separated; - the detection on said history of a single characteristic point in the portion of the history corresponding to area 1, area 4 and the interface between these two areas; - the comparison of the position of the characteristic point against a target position; - the adjustment of the carrying volume of area 1 and / or the carrying volume of area 4, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position; - the volume of the mobile phase injected per cycle being maintained greater than, or equal to, a minimum limit and / or less than, or equal to, a maximum limit.

2. Method according to claim 1, wherein the volume of mobile phase injected per cycle is maintained at a constant value.

3. Method according to claim 1 or 2, wherein the target position of the characteristic point is defined relative to the position of the step of injecting the mobile phase and / or to the position of the step of collecting the extract and / or to the position of the step of collecting the raffinate.

4. Method according to any one of claims 1 to 3, wherein the variable that is representative of the concentration of one or more species contained in the mixture to be separated, is chosen from optical rotation, absorbance, emission of spectroscopic radiation, refractive index, density, conductivity, pH and combinations thereof.

5. Method according to any one of claims 1 to 4, wherein the characteristic point is chosen from a local minimum of the history, a point of the history having a determined value, and a barycenter of two points of the history corresponding to values defined absolutely or relatively.

6. Method according to any one of claims 1 to 5, further comprising the steps of: - the measurement of the purity and / or of the yield of at least one collected fraction chosen from the extract and the raffinate; - the comparison of the measured purity and / or the measured yield against a target purity and / or a target yield; and optionally comprising the steps of: - the measurement of the purity and / or the yield of the raffinate and of the purity and / or the yield of the extract; - the comparison of the measured purity and / or of the measured yield of each fraction with, respectively, a target purity and / or a target yield.

7. Method according to claim 6, further comprising: - a step of modifying the volume of mixture to be separated that is injected per cycle according to the difference between the measured purity(ies) and / or yield(s), and the target purity(ies) and / or yield(s); or - a step of modifying the volume of mobile phase injected per cycle according to the difference between the measured purity(ies) and / or yield(s) and the target purity(ies) and / or yield(s); or - a step of jointly modifying the volume of mixture injected per cycle and the volume of mobile phase injected per cycle according to the difference between the measured purity(ies) and / or yields and the target purity(ies) and / or yield(s).

8. Method according to claim 6 or 7, further comprising a step of defining the target position according to the difference between the measured purity(ies) and / or yield(s) and the target purity(ies) and / or yield(s).

9. Method according to any one of claims 1 to 8, further comprising: - the determination, in a node of the system, of the history of a variable that is representative of the concentration of one or more species contained in the mixture to be separated; - the detection on the history of another characteristic point in the portion of the history corresponding to area 2, area 3 and the interface between these two areas; - the comparison of the position of the other characteristic point against a target position; - the adjustment of the carrying volume of area 2 and / or the carrying volume of area 3, modifying the position of the other characteristic point to bring it closer to its target position.

10. Method according to any one of claims 1 to 9, wherein the injection of the mixture to be separated is a continuous flow and, preferably, the method is a simulated moving bed method; or wherein the injection of the mixture to be separated is discontinuous, and, preferably, the method is a sequential simulated moving bed method.

11. Method according to any one of claims 1 to 10, wherein the mixture to be separated comprises one or more monosaccharides, preferably glucose and fructose, and wherein the extract and the raffinate are enriched with different monosaccharides.

12. Method according to any one of claims 1 to 11, wherein: the volume of the mobile phase injected per cycle is maintained greater than, or equal to, a minimum limit and / or less than, or equal to, a maximum limit, and the method comprises: - the determination, in a node of the system, of the history of a variable that is representative of the concentration of one or more species contained in the mixture to be separated; the method being such that: - as long as the volume of the mobile phase injected per cycle is greater than, or equal to, said minimum limit, or less than, or equal to, said maximum limit, the method comprises: ▪ the detection on the history of a first characteristic point in the portion of the history corresponding to area 1; ▪ the detection on the history of a second characteristic point in the portion of the history corresponding to area 4; ▪ the comparison of the position of each of the characteristic points against a respective target position; ▪ the adjustment of the carrying volume of area 1 and the carrying volume of area 4, modifying the position of the first characteristic point and the second characteristic point, respectively, to bring them closer to their respective target positions; - when the volume of the mobile phase injected per cycle reaches the maximum or the minimum limit, the method comprises: ▪ the detection on said history of a single characteristic point in the portion of the history corresponding to area 1, area 4 and the interface between these two areas; ▪ the comparison of the position of the characteristic point against a target position; ▪ the adjustment of the carrying volume of area 1 and / or the carrying volume of area 4, modifying the position of the characteristic point to bring the position of the characteristic point closer to the target position.

13. Computer program comprising program code instructions for the execution of the steps of the method according to any one of claims 1 to 12 when said program is executed on a computer.

14. Computer-readable storage medium on which a computer program according to claim 13 is recorded, or system comprising a processor coupled to a memory on which a computer program according to claim 13 is recorded.