DESC:TECHNICAL FIELD OF THE INVENTION
The present invention relates to a process for separating and purifying sugar component from the ternary mixture comprising of monosaccharides and/or disaccharides using continuous ternary separation chromatography system with variable length chromatography zones. The present invention particularly relates to a process for production of allulose or sucrose isomers using ternary separation chromatography system with variable length chromatography zones. The present invention is an improvement over the conventional method which fails to separate ternary sugar mixtures comprising of monosaccharaides and/ or disaccharides.

BACKGROUND OF THE INVENTION
Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis. There are several types of chromatography, affinity chromatography, ion exchange chromatography, size exclusion chromatography or reversed phase chromatography among others.
Lab scale chromatographic purifications are carried out in a batch mode, where a single column is loaded with the component mixture to be separated followed by elution with a mobile phase. Batch mode operation for large volumes or purifying large amounts of product would hence require large column volumes and multiple injections or several columns operating in parallel. This leads to commercially unviable option.
The limitations in the batch mode chromatographic separation led to the development of continuous chromatography systems for industrial application. These continuous chromatography systems are based on multiple columns. In 1961, Broughton et al. of UOP proposed the concept of simulated moving bed (SMB) for the separation of xylene isomers in the petrochemical area. With a wide range of commercial applications, simulated moving bed (SMB) chromatography has emerged as an extremely effective adsorption-based separation technique. The biochemical and pharmaceutical industries have effectively incorporated its use. Due to its effective separation mechanism and continuous operating mode, SMB offers high product purity and yield. Subsequently, it was progressively implemented in the sugar business and chiral drug resolution fields.
The True Moving Bed (TMB), serves as the foundation for the development of the standard SMB system.
Several novel SMB modes have been put forth. For instance, the Ludemann-Hombourger-proposed VariCol system, which is based on non-synchronous switching of inlet/outlet ports, enhances SMB performance within a cycle, increasing operational flexibility and requiring a much lesser number of columns.
US patent 6,712,973 B2 discloses a process and device for the separation of components with variable chromatographic zones. However, this disclosure cannot be used for separation of ternary mixtures.
Allulose, a rare sugar, also known as D-psicose is a low-calorie natural sweetener that has 70% the sweetness of sucrose, but only 10% of the calories. It is a naturally occurring monosaccharide and present in small quantities in wheat and other plants, hence also classified under “Rare Sugars”. It has several health benefits: close to zero calorie, very low glycemic index of 1, it helps regulate the blood sugar etc.
At present, D-allulose is produced from fructose using D-allulose 3-epimerase as the catalyst. The substrate conversion of this enzymatic conversion is low and the highest conversion is only 32%. This process requires a simulated moving bed chromatography system to realize the recycling of fructose which increases the production cost. Therefore, there is a need for an alternative cost-effective approach for high yield allulose production.
US patent application 2023/0046104 discloses a process for the production of psicose (allulose) from fructose raw material. In the disclosure a SMB has been used for separating the binary mixture of fructose and allulose. The limitation of this process is that the SMB system cannot be used for the separation of ternary mixtures and hence other low-cost raw materials like starch cannot be used for the production of allulose.
US patent application 2023/0058087A1 discloses a process for production of allulose. According to the disclosure the separation of sugars from a mixture containing allulose, glucose and fructose is achieved using a conventional simulated moving bed chromatography system. The conventional SMB used in US’087 employs a 12 column SMB system wherein the resin required is very high that adds to the cost of the sytem. Also, when the process is carried out with other raw materials like starch in US’087an additional SMB is required in the fructose refining section. Moreover, US’087 disclose the production using HFCS only as the feed material. Furthermore, though US’087 disclose high yield of allulose, the productivity i.e. the amount of allulose recovered per unit volume of resin per hour low which is usually expressed in g/l/h is low since US’087 uses 12 columns the resin used would be high thereby decreasing the productivity.
Therefore, there is need for the development of an efficient, cost effective and simple system for separating components from the sugar mixture comprising of more than two components.
Patent application US 2019/0330253 A1, describes a process to produce allulose from a fructose solution. The process employs a conventional simulated moving bed chromatography system for binary sugar separation and at-least one nanofiltration system to produce allulose crystals. The scale up of such a process to industrial scale would not be economically viable.
Patent application US 2023/0058087 A1, describes a process to produce allulose from high fructose corn syrup. The process utilizes a conventional simulated moving bed chromatography system for separation of ternary sugar mixture comprised of glucose, fructose and allulose.
The process for the production for allulose in the prior art utilize fructose primarily as the raw material. In light of the same, the present inventors felt that there is a scope for production of allulose or sucrose isomers from other cheap sugar sources such as starch using continuous ternary separation chromatography system with variable length chromatography zones.

OBJECT OF THE INVENTION
The primary object of the present invention is to provide a process for separating and purifying sugar component such as allulose or sucrose isomer such as isomaltulose, trehalulose from the ternary mixture using continuous ternary separation chromatography system with variable length chromatography zones in a cost effective manner.

SUMMARY OF THE INVENTION:
In accordance with the above, the present invention provides a process for separating and purifying the sugar component(s) from the ternary mixture using continuous ternary separation chromatography system with variable length chromatography zones comprising;
i. introducing a feed solution containing a ternary mixture selected from monosaccharides and/or disaccharides into at least one of the chromatography columns (C1-C6) having a chromatographic bed material through its corresponding feed pump;
ii. receiving the diluent/eluent into at least one of the chromatography columns (C1-C6) through its corresponding diluent pump;
iii. continuously withdrawing an extract fraction from the chromatography columns (C1-C6) through its corresponding extract pump,
iv. continuously withdrawing the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) through its corresponding raffinate pump;
wherein said chromatography system comprises plurality of columns positioned adjacent to each other and fluidically connected in series forming a closed loop configuration and configured to operate with variable length chromatography zones by asynchronous switching of its inlet and outlet ports.
In an aspect, the feed solution containing a ternary mixture preferably comprises of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose. More preferably, the feed solution containing a ternary mixture comprises of allulose, one or more isomers, including epimers of allulose and one or more additional sugars like glucose and its isomers.
In yet another aspect the diluent is preferably water.
In another aspect, the chromatographic bed material could be selected based on its differential adsorption characteristics towards the sugars in the feed solution preferably the suitable chromatographic bed materials include but are not limited to resins that are strong acid cation resins. Some examples of suitable strong acid cation resins include product sold by Dow Chemicals Co (Dowex99 Ca/320; Dowex99 Ca/310 and Dowex99 Ca/280), Mitsubishi Chemical Co. (UBK 555) and Purolite (PCR642 Ca).
In another aspect, said continuous ternary separation chromatography system is configured to operate in sub-cycles comprising;
i. Sub-cycle 1 wherein at time (t) providing the ternary mixture to column 1 (C1) and simultaneously drawing off the raffinate from column 2 (C2) and simultaneously providing the diluent to column 4 (C4) and drawing-off the extract from column 4 (C4) resulting in zone I comprising of one column length, zones II and III comprising of two columns each and zone IV comprising of a single column;
ii. Sub-cycle 2 wherein after a certain point in time (t+Td2) shifting the extract drawing-off point alone by one column from column 4 (C4) to column 5 (C5) while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone III comprises of two columns and zone II and zone IV comprising of single column;
iii. Sub-cycle 3 wherein after a certain point in time (t+Td3) shifting the eluent inlet to column 5 (C5) from column 4 (C4), while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone II comprises of one column and zone III and zone IV comprising of two columns;
iv. Sub-cycle 4 wherein after a certain point in time (t+Td4) shifting the feed inlet from column 1 (C1) to column 2 (C2), while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone III comprises one column and zone II and zone IV comprising two columns;
wherein after a certain point in time, initiating cycle 2 by one column increment at inlet and outlet valve positions and the sub-cycles 2, 3, and 4 follow the same pattern as in cycle 1; and
continuing the operation following the above scheme for an extended number of cycles for continuous separation of components.
In an aspect, a process for production of D-allulose comprising;
i. Section 20 comprising systems to generate an aqueous solution of raw material selected from monosaccharides, dissacharides, polysaccharides, oligosaccharides or mixtures thereof;
ii. Section 30 comprising of epimerization and refining systems for converting a part of the raw material from section 20 to an aqueous solution of ternary mixture of isomerized or epimerized sugars comprising of primarily monosaccharides without any ionic impurities;
iii. Section 40 comprising of a continuous ternary separation chromatography system of the present invention for separation and purification of desired product from the ternary sugar mixture, wherein the mixture of unreacted sugars as stream 1 and stream 2 are recycled to section 20 and the desired product as stream 3 is further processed in section 50;
iv. Section 50 comprising of systems for the downstream processing of the desired product to obtain in crystal form comprising the concentration systems, demineralization and decolorization systems, crystallization systems, solid-liquid separation systems, and drying systems among others; and recycling the mother liquor obtained after the crystallization step to section 30 to increase the overall conversion of the raw material for continuous separation and purification of the desired component.
The above sections are the parts of the process to obtain the sugar component. The Varicol system is one section in the sugar component production process.
In an aspect, the process of the present invention may be carried out in a ternary chromatographic system comprising of 6 columns and 4 zones.
In yet another aspect, the present invention provides a continuous process for production of D-allulose from saccharides or sugar mixtures with unique and advantageous recycle schemes, including an efficient multi-column chromatography separation system wherein a mixture comprising of more than two components are separated by contact between solid and liquid phases in variable length chromatography zones.
The saccharides or sugar mixtures that can be used to produce the D-allulose is selected from starch and its derivatives like amylose, amylopectin, maltodextrin, maltose, fructose and glucose, cellulose derivatives which include pretreated biomass, amorphous cellulose, cellodextrin, cellobiose and glucose, sucrose derivatives include glucose and fructose.
The derivatives of starch and starch can be utilized after enzymatic hydrolysis or acid hydrolysis of starch and/or its derivatives. The derivatives of cellulose and cellulose can be utilized after the enzymatic hydrolysis of cellulose and/ or its derivatives in section 20. The hydrolysis step may also be accomplished by acid hydrolysis with or without the pretreatment of biomass. Sucrose derivatives, glucose and fructose, can be produced by enzymatic hydrolysis using invertase enzyme in free or immobilized form or combination thereof or hydrolysis using strong acid cation resins or other acid hydrolysis process or their combinations thereof.
In an aspect, the present invention provides a process for production of D-allulose comprising;
i. generating an aqueous solution of monosaccharides selected from glucose and fructose by enzymatic hydrolysis or acid hydrolysis or isomerization of the saccharides or sugar mixtures followed by concentrating in section 20;
ii. epimerizing the aqueous solution rich in monosaccharaides to obtain glucose, fructose and D-allulose and refining to remove ionic impurities in section 30;
iii. separating the glucose, fructose and D-allulose ternary sugar mixture in section 40 comprising the continuous ternary separation chromatography system with variable length chromatography zones of the present invention configured to operate in sub-cycles resulting in a first process stream comprising an aqueous solution rich in glucose; a second process stream comprising a mixture of glucose and fructose; and a third process stream comprising an aqueous solution rich in D-allulose;
iv. recycling the first process stream rich in glucose to the isomerization step in zone 20 to obtain the mixture of glucose and fructose;
v. mixing the second process stream with glucose and fructose of step (iv);
vi. subjecting the D-allulose obtained in third process stream to downstream process consisting of concentration(s), demineralization, decolorization, crystallization(s), solid-liquid separation and drying;
vii. recycling a portion of the mother liquor obtained after crystallization of step (vi) to the step of converting a portion of the fructose to D-allulose;
viii. recycling a portion of the mother liquor obtained after the crystallization step (vi) to the continuous ternary separation chromatography system to obtain D-allulose rich fraction.
In an aspect of the invention, the continuous ternary separation chromatography system, results in three streams, an aqueous stream rich in glucose, an aqueous stream rich in glucose and fructose with negligible D-allulose and an aqueous stream rich in D-allulose. The glucose rich fraction is recycled to an isomerization step in section 20 and result in a mixture of glucose and fructose. The requirement of a concentration step is avoided resulting reduction of capital and operating costs.
In another aspect, the aqueous stream rich in glucose and fructose with negligible D-allulose obtained after the continuous ternary separation chromatography is recycled and mixed with the glucose and fructose fraction obtained after the isomerization step. This fraction is directly used as the feed for the epimerization step after a concentration step. This operation mode avoids the requirement of an additional chromatographic separation process to generate fructose rich fraction for the epimerization step. The mixing of the aqueous stream rich in glucose and fructose with negligible D-allulose obtained after the continuous ternary separation chromatography is recycled with the glucose and fructose fraction obtained after the isomerization step also increases the total sugar concentration to the concentration step and results in reduced energy for the concentration step.
In another aspect, the mother liquor obtained after the crystallization step is recycled to the continuous ternary separation chromatography system which increases the overall conversion of the raw material to crystal D-allulose.
In yet another aspect, the present invention provides a process for separation and purification of sucrose isomers from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose using the continuous ternary separation chromatography system wherein the chromatography system is configured to continuously withdraw an extract fraction from at least one of the chromatography columns (C1-C6) by its corresponding extract pump, wherein the extract fraction comprise of an aqueous solution rich in first sucrose isomer preferably isomaltulose.
In another aspect, the present invention provides a process for separation and purification of sucrose isomer from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose using the continuous ternary separation chromatography system wherein the chromatography system is configured to continuously withdraw the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) by its corresponding raffinate pump, wherein the raffinate 1 fraction comprise of an aqueous solution rich in second sucrose isomer preferably trehalulose and raffinate 2 comprising of a mixture of glucose and fructose.
These and other aspects, embodiments, and associated advantages will become apparent from the following brief description of the drawings and the detailed description.
BRIEF DESCIRPTION OF DRAWINGS
Figure 1: Schematic of conventional SMB operation
Figure 2: Schematic of operation with variable length chromatography zones.
Figure 3: Schematic flow diagram of the system, with 6 columns in accordance with certain aspects of the present invention.
Figure 4: Schematic view depicting a process of producing D-allulose according to
DETAILED DESCRIPTION OF THE INVENTION
The foregoing descriptions of specific embodiments of the present invention have been presented for the purposes of illustration and description. They are not intended to be exhaustive or to limit the invention to the precise forms disclosed, and obviously many modifications and variations are possible in light of the above teaching.

Unless specified otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art, to which this invention belongs. Although any process and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred process and materials are described.

Unless stated to the contrary, any of the words “contains”, “containing”, "including," "includes," "comprising," and "comprises" mean "including without limitation" and shall not be construed to limit any general statement that it follows to the specific or similar items or matters immediately following it. Embodiments of the invention are not mutually exclusive but may be implemented in various combinations. The described embodiments of the invention and the disclosed examples are given for the purpose of illustration rather than limitation of the invention.

Further, words like “a”, “an”, “at least” and “the” should be construed to not only cover singular quantities but also plural quantities of the elements immediately following them.

The term “Chromatographic bed material” refers to a solid phase sorbent material used to separate the chemical species by sorbent separation.

The term “eluent/diluent” used interchangeably in the entire specification refers to the mobile phase passed over the chromatographic bed to achieve sorbent separation.
The term “raffinate” refers to the liquid effluent or fraction resulting from a separation process not enriched with the desired sugar component.

The term “non-constant time” as used herein in the entire specification means and relates to situations where the time taken to complete an operation or function varies depending on the parameters of operation.
The term “column increment” as used herein in the entire specification means and relates to the periodic switching or shifting of inlet and outlet ports to simulate the movement of the solid phase (stationary phase), even though the columns themselves do not physically move.
The term “chromatography system” or “system” used interchangeably in the entire specification means and refer to the continuous ternary separation chromatography system with variable length chromatography zones.
Abbreviations:
HWS (in figure 4)-Hot water supply
HWR (in figure 4)-Hot water return.
The present invention relates to a process for separating and purifying sugar component(s) from the ternary mixture using continuous ternary separation chromatography system with variable length chromatography zones. While the prior arts describe the simulated moving bed comprising of plurality of column segments sequentially connected in series and containing the chromatography beds for separation of sugar, the process disclosed therein are costly due to the use of high amount of resins. Moreover, the processes disclosed therein use a single raw material preferably fructose source. The present invention provides a process for separating and purifying sugar component(s) using the continuous ternary separation chromatography system with less number of columns and from a raw material selected from monosaccharides, disaccharides, polysaccharides, oligosaccharides or mixtures thereof.
Prior to describing the present invention in detail, the principle of continuous chromatography for separation of components in variable length chromatography zones shall be detailed.
There are different chromatographic processes that can be used for the separation of chemical component on a large scale. One such possibility is the true standard moving bed 4-zone counter current process which is a simulated moving bed system. In this system, solids circulate continuously in a closed loop past the fixed feed and eluent introduction points alternately with draw off points for a raffinate and an extract. The system can be divided into four different zones:
Zone 1: everything located between the eluent and extract lines;
Zone 2: everything between the extract and feed lines;
Zone 3: everything between the feed and raffinate lines;
Zone 4: everything between the raffinate and eluent lines.
In a true moving bed, the flow of the solids is constant throughout the system. However, in view of the inlet/ outlet flow rates, the liquid flow rate varies depending on the zone. Accordingly, QI, QII, QIII and QIV are the respective flow rates in the zones I, II, III and IV.
The simulated moving bed (SMB) chromatography makes it possible to simulate the movement of the solid using the connection between the columns that are interconnected in a closed loop. The SMB operates by shifting the inlet and the outlet points at fixed intervals in a multi-column system. As illustrated in Figure 1, the feed stream containing weakly adsorbed (light) and strongly adsorbed (heavy) components enters the system between zones II and III. Separation is achieved on switching the columns as heavy component remains in zone II with the solid phase while light component is desorbed by the eluent and enters into zone III with the liquid phase. The operating parameters are adjusted such that the two components move counter current to each other to achieve separation. Thus, zone II and zone III are referred as the separation zone. After that, heavy components are desorbed in zone I to regenerate the solid phase. Hence, zone I is also called as solid phase regeneration zone. Zone IV is termed as the liquid phase regeneration zone, which adsorbs the light component and regenerates the liquid phase.
The following are the main characteristics of the SMB:
• The zones are defined by the position of the inlet/outlet
• The number columns per zone are fixed
• The length of the zones is hence fixed
• There is synchronized switching of all the inlet/outlet.
With reference to Figure 2, separations with variable length chromatography zones (VariCol) are carried out by asynchronous switching mode. In VariCol, every cycle time is divided into sub-cycles as represented in Table 1. In each sub-cycle the zone lengths are not fixed but are varied over time. The zone lengths oscillate continually by one column, with the decrease of one zone offsetting the increase of the adjacent zone and vice versa. The VariCol mode of operation of the present invention using six column assemblies is shown in Figure 2.
Table 1 - VariCol configuration for 6 column arrangement corresponding to first cycle
Sub-cycle Time Inlet/Outlet No. of columns in each zone (I/II/III/IV)
1 t + dT1 D(4)/E(4)/F(1)/R(2) 1/2/2/1
2 t + dT2 D(4)/E(5)/F(1)/R(2) 2/1/2/1
3 t + dT3 D(5)/E(5)/F(1)/R(2) 1/1/2/2
4 t + ?T D(5)/E(5)/F(2)/R(2) 1/2/1/2
Average no. of columns 1.25/1.5/1.75/1.5

The Table 1 corresponds to first cycle with feed inlet to column 1. Next cycle starts at the end of sub-cycle 4 of cycle 1 with inlet/outlet positions change by one column length as D(5)/E(5)/F(2)/R(3).
The present invention provides advantages over conventional processes and products. The present invention provides an efficient multi-column chromatography separation process for separation of ternary mixtures with variable length chromatography zones (VariCol).
In an embodiment, the present invention relates to a system for purifying sugars from a ternary mixture, the system comprises of a continuous ternary chromatography system having a plurality of columns positioned adjacent to each other and fluidically connected in series forming a closed loop configuration, wherein each column in the system is defined with a feed pump, diluent pump, extract pump, and raffinate pump. The continuous ternary chromatography system for carrying out the separation and purification of sugar component(s) from the ternary mixture is illustrated in Fig 3.
In an embodiment, the process for separating and purifying the sugar component(s) from the ternary mixture using continuous ternary separation chromatography system with variable length chromatography zones comprising;
i. introducing a feed solution containing a ternary mixture selected from monosaccharaides and/or disaccharides into at least one of the continuous ternary separation chromatography columns (C1 to C6) having a chromatographic bed material through its corresponding feed pump;
ii. receiving the diluent/eluent into at least one of the chromatography columns (C1 to C6) through its corresponding diluent pump;
iii. continuously withdrawing an extract fraction from the chromatography columns (C1 to C6) through its corresponding extract pump,
iv. continuously withdrawing the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1 to C6) through its corresponding raffinate pump and
wherein said chromatography system comprises the columns (C1-C6) positioned adjacent to each other and fluidically connected in series forming a closed loop configuration and configured to operate with variable length chromatography zones by asynchronous switching of its inlet and outlet pumps.
Accordingly, said system comprises of a plurality of columns (C1-C6) that contain solid bed material arranged in series and in a closed loop, whereby said loop comprises a number of fluid injection lines in each column or column section that are connected to at least one injection pump and a number of fluid draw-off lines of each column or column section that are connected to at least one draw off means, at least one valve on each line, whereby said loop defines at least three chromatographic zones, whereby each of this is determined by a fluid injection point and a fluid draw-off point, whereby the system is characterized in that it comprises means for controlling the variation in time of the length of the zones that are connected to said valve and that are suitable for shifting by one column or column section the positions of the injection points and draw-off points in an intermittent manner.
In an embodiment, the continuous chromatography system is configured to receive the feed solution containing a ternary mixture into at least one of the chromatography columns (C1-C6) through its corresponding feed pump (3) and wherein the feed solution contains a ternary mixture comprising of monosaccharides and/or disaccharides. Preferably, the feed solution containing a ternary mixture comprises of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose. More preferably, the feed solution containing a ternary mixture comprises of allulose, one or more isomers including epimers of allulose and one or more additional sugars like glucose and its isomers.
In yet another embodiment the diluent/eluent used is preferably water.
In yet another embodiment, the extract fraction from at least one of the chromatography columns is continuoulsy withdrawn by its corresponding extract pump, wherein the extract fraction may comprise of an aqueous solution rich in allulose or first sucrose isomer such as isomaltulose or trehalulose.
In another embodiment, in the process of the present invention the raffinate 1 or raffinate 2 are continuously withdrawn from at least one of the chromatography columns by its corresponding raffinate pump, wherein the raffinate 1 fraction may comprise of an aqueous solution rich in glucose or a second sucrose isomer and raffinate 2 comprises of a mixture of glucose and fructose.
In an embodiemnt, the shifting of the positions of the injection points and draw-off points is in the same direction as that of the flow in the columns.
In yet another embodiment, the flow rate of the fluid that circulates in a given zone is generally constant.
In another embodiment, during the cycle time the position of the injection or draw-off points is shifted with non-constant time shift.
In an embodiment, the present invention discloses a process, during the cycle time where it is possible to shift all the injection and draw-off positions with a constant time and advantageously with a time phase shift that is at-least equal to a quarter cycle time.
In another embodiment, the means of draw off at the extract and raffinate lines is by using a pump. Alternatively, an analog valve connected to a flow meter can also be used.
In an embodiment, the columns in the continuous chromatography system are filled with the chromatographic bed material, wherein the bed material is an ion exchange resin. Examples of suitable chromatographic bed materials include but not limited to, resins that are strong acid cation resins. Some examples of suitable strong acid cation resins include product sold by Dow Chemicals Co (Dowex99 Ca/320; Dowex99 Ca/310 and Dowex99 Ca/280), Mitsubishi Chemical Co. (UBK 555) and Purolite (PCR642 Ca).
In yet another embodiment, the present invention relates to a process for separation and purification of allulose from a sugar mixture comprising of glucose, fructose and allulose wherein the continuous chromatography system is operated as per the scheme provided in Table 1 - VariCol configuration for 6 column arrangement corresponding to first cycle. The process comprises the steps of:
i. Sub-cycle 1: Providing the feed sugar mixture comprising of glucose, fructose and allulose to column 1 (C1) and simultaneously drawing off the raffinate 1 or 2 from column 2 (C2) and simultaneously providing the eluent to column 4 (C4) and drawing-off the extract from column 4 (C4) resulting in zone I comprising of one column length, zones II and III comprising of two columns each and zone IV comprising of a single column;
ii. Sub-cycle 2: After a certain point in time (t+Td2) advantageously shifting the extract drawing-off point alone by one column from column 4 (C4) to column 5 (C5) while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are comprised of two columns and zone II and zone IV are comprised of single column;.
iii. Sub-cycle 3: After a certain point in time (t+Td3) advantageously shifting the eluent inlet to column 5 (C5) from column 4 (C4), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone II are now comprised of one column and zone III and zone IV are comprised of two columns;
iv. Sub-cycle 4: After a certain point in time (t+Td4) advantageously shifting the feed inlet from column 1 (C1) to column 2 (C2), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are comprised on one column and zone II and zone IV are comprised on two columns. The four steps correspond to cycle 1;
wherein after a certain point in time, initiating cycle 2 by one column increment at inlet and outlet valve positions and the sub-cycles 2, 3, and 4 follow the same pattern as in cycle 1; and
continuing the operation following the above scheme for an extended number of cycles for continuous separation of components.
In another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose wherein the continuous chromatography sytsem is configured to continously withdraw an extract fraction from at least one of the chromatography columns (C1-C6) by its corresponding extract pump, wherein the extract fraction may comprise of an aqueous solution rich in allulose.
In yet another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose, wherein the continuous chromatography system is configured to continously withdraw the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) by its corresponding raffinate pump , wherein the raffinate 1 fraction may comprise of an aqueous solution rich in glucose and raffinate 2 comprises of a mixture of glucose and fructose.
In an embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose in various concentrations wherein the concentration of glucose, fructose and allulose are preferably in the range of 10 – 40%, 15 – 40% and 1 – 25% respectively.
In another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose wherein cycle switching time is in between 40-120 min.
In yet another embodiment of the present invention, the feed stream has a flow rate of about 10 cm/h to 60 cm/h, the eluent has a flow rate ranging from 60 cm/h to 140 cm/h, the extract has a flow rate ranging from 20 cm/h to 90 cm/h, the raffinate has a flow rate ranging from 30 cm/h to 110 cm/h.
According to another embodiment of the present invention, the temperature of the system is maintained between 30°C – 90 °C.
In certain embodiments, the present invention relates to a process and system for separation and purification of sugars from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose wherein the continuous chromatography system is operated as per the scheme provided in Table 1 - VariCol configuration for 6 column arrangement corresponding to first cycle. The process comprises the steps of:
i. Sub-cycle 1: Providing the feed sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose to column 1 (C1) and simultaneously drawing off the raffinate 1 or 2 from column 2 (C2) and simultaneously providing the eluent to column 4 (C4) and drawing-off the extract from column 4 (C4) resulting in zone I comprising of one column length, zones II and III comprising of two columns each and zone IV comprising of a single column;
ii. Sub-cycle 2: After a certain point in time (t+Td2) advantageously shifting the extract drawing-off point alone by one column from column 4 (C4) to column 5 (C5) while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are now comprised of two columns and zone II and zone IV are comprised of single column;
iii. Sub-cycle 3: After a certain point in time (t+Td3) advantageously shifting the eluent inlet to column 5 (C5) from column 4 (C4), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone II are now comprised of one column and zone III and zone IV are comprised of two columns;
iv. Sub-cycle 4: After a certain point in time (t+Td4) advantageously shifting the feed inlet from column 1 (C1) to column 2 (C2), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are comprised on one column and zone II and zone IV are comprised on two columns. The four steps correspond to cycle 1;
wherein after a certain point in time, initiating cycle 2 by one column increment at inlet and outlet valve positions and the sub-cycles 2, 3, and 4 follow the same pattern as in cycle 1; and
continuing the operation following the above scheme for an extended number of cycles, for continuous separation of components.
In yet another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose wherein the continuous chromatography sytsem is configured to continously withdraw an extract fraction from at least one of the chromatography columns (C1-C6) by its corresponding extract pump, wherein the extract fraction may comprise of an aqueous solution rich in first sucrose isomer or more preferably isomaltulose.
In yet another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose wherein the continuous chromatography system is configured to continously withdraw the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) by its corresponding raffinate pump (8), wherein the raffinate 1 fraction may comprise of an aqueous solution rich in second sucrose isomer or more preferably trehalulose and raffinate 2 comprises of a mixture of glucose and fructose.
In yet another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose wherein the concentration of first sucrose isomer or isomaltulose, second sucrose isomer or trehalulose and other sugars including monosaccharides are preferably in the range of 30 – 90%, 3 – 30% and 2 – 30% respectively.
In another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose wherein cycle switching time is in between 50-120 min.
In yet another embodiment, the feed stream has a flow rate about 10 cm/h to 60 cm/h, the eluent has a flow rate ranging from 60 cm/h to 140 cm/h, the extract has a flow rate ranging from 20 cm/h to 90 cm/h, the raffinate has a flow rate ranging from 30 cm/h to 110 cm/h.
In another embodiment, the temperature of the system is maintained at a temperature ranging between 30°C – 90 °C.
According to another embodiment of the present invention, the number of columns or column sections is temporarily zero in one or more zones of the system
In an embodiment, for a given total column number infinite possible configurations with the VariCol process which depend on the phase shift of the switching are carried out.
The present invention thus provides a continuous process for production of D-allulose or sucrose isomers such as isomaltulose or trehalulose from sugar mixtures with unique and advantageous recycle schemes, including an efficient multi-column chromatography separation system wherein a mixture comprising of more than two components are separated by contact between solid and liquid phases in variable length chromatography zones.
In an embodiment, the Figure 14 of the present invention depicts a flow diagram illustrating production of allulose from sucrose and/ or high fructose syrups and/or high fructose corn syrups and/ or fructose containing syrups and/ or glucose syrups and/ or polysaccharides like starch and cellulose which comprises of the following sections:
Section 20: comprising systems to generate an aqueous solution rich in monosaccharides, wherein the monosaccharides are primarily glucose and fructose.
Section 30: comprising of systems for converting part of the fructose in the monosaccharide sugar solution from section 20, comprising primarily of glucose and fructose, to D-allulose. The conversion system utilizes a catalyst more preferably a biological catalyst and results in an aqueous solution comprising of glucose, fructose and D-allulose.
Section 40: comprises of system for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and D-allulose. The system comprising of a continuous ternary separation chromatography system of the present invention having a plurality of columns positioned adjacent to each other and fluidically connected in series forming a closed loop configuration. The continuous ternary separation chromatography system is operated with variable length chromatography zones results in three process streams, stream 1 comprises of aqueous solution rich in glucose and stream 2 comprises of a mixture of glucose and fructose and finally stream 3 comprises of an aqueous solution rich in D-allulose. The streams 1 and 2 are advantageously recycled to section 20 and stream 3 is further processed in section 50
Section 50: comprises of systems for the downstream process of D-allulose required for producing D-allulose crystals. The systems comprise of concentration systems, demineralization and decolorization systems, crystallization systems, solid-liquid separation systems, and drying systems among others. The mother liquor obtained after the crystallization step is advantageously recycled to section 30 to increase the overall conversion of the raw material to crystal D-allulose.
As shown in section 20 of Figure 4 the process for the production of aqueous solution rich in monosaccharides, primarily glucose and fructose, utilizes sucrose and/ or high fructose syrups and/or high fructose corn syrups and/ or fructose containing syrups and/ or glucose syrups and/ or polysaccharides like starch and cellulose and/or sugar containing solutions.
As shown in section 20 of Figure 4 the process for the production of an aqueous solution rich in monosaccharides, primarily glucose and fructose, and/or sucrose, may comprise of remelt system, inversion system and refining system 1. The remelt system may comprise of an agitated vessel to which sucrose and demineralized water are added to result in final concentration of sucrose of not less than 25%. The remelt process may be carried out at ambient temperature or at a temperature between 30 to 90°C. In certain aspect, the remelt process may be carried out using a continuous sugar dissolution system.
In another embodiment, the inversion system, which results in the hydrolysis of sucrose to aqueous mixture of sugars primarily composed of glucose and fructose may be comprised of a single or plurality of columns containing immobilized enzyme or solid acid catalyst or strong acid cation resins. In some aspects, the columns may be operated in series or in parallel configuration to result in near 100% hydrolysis of sucrose to an aqueous mixture of sugars primarily composed of glucose and fructose. In certain aspects, the inversion system, may be comprised of a single or plurality of stirred tank reactors operating in parallel to which food grade acids or inversion enzyme is added to hydrolyze sucrose to aqueous mixture of sugars primarily composed of glucose and fructose. In another aspect, the inversion system may be operated at a temperature above ambient, preferably at a temperature between 30°C and 90°C, more preferably above 50 °C.
In another embodiment of the present invention, as shown in section 20 of Figure 4, the saccharides that can be used for the production of D-allulose, comprises of polysaccharides, oligosaccharides, and/or disaccharides. The saccharides can be starch and its derivatives like amylose, amylopectin, maltodextrin, maltose, fructose and glucose. The derivatives of starch and starch can be utilized after enzymatic hydrolysis or acid hydrolysis or other processes know in the art. In this aspect, the solution obtained after the enzymatic hydrolysis of starch and its derivatives or by acid hydrolysis of starch and its derivatives comprising primarily glucose is subjected to an isomerization step, wherein the glucose present in the solution is converted to aqueous mixture of sugars primarily composed of glucose and fructose. In this aspect, the isomerization system may comprise of immobilized enzymes in packed bed reactor systems or free enzymes in solution in a stirred tank reactor. The system comprises of single reactor of any of the said reactor types or plurality of reactors of any of the said reactor types arranged in series or in parallel. In certain aspects, the enzyme is immobilized onto the solid matrix by means of adsorption or gel entrapment or covalent cross linking or any other process known in art. In certain aspects, the residence time of the feed solution in the different reactors of the isomerization system may be uniform or different. In some aspects, the reaction in the isomerization system is carried out at a pH between 5.0 to 9.0 and conducted at a temperature between 35 – 90 °C by using a suitable isomerase enzyme. Such isomerase includes enzymes but not limited to those involved the conversion of glucose to fructose.
In another embodiment of the present invention, as shown in section 20 of Figure 1, the saccharides that can be used for the production of the D-allulose include cellulose and its cellulose derivatives. In this aspect, the saccharides include pretreated biomass, amorphous cellulose, cellodextrin, cellobiose and glucose. The derivatives of cellulose and cellulose can be utilized after the enzymatic hydrolysis using a mixture of cellulose enzymes, alternatively they can be utilized after acid hydrolysis with or without the pretreatment. In this aspect, the solution obtained after the enzymatic hydrolysis of cellulose and its derivatives or by acid hydrolysis of cellulose and its derivatives comprising primarily glucose is subjected to an isomerization step, wherein the glucose present in the solution is converted to aqueous mixture of sugars primarily composed of glucose and fructose. In this aspect, the isomerization system may comprise of immobilized enzymes in packed bed reactor systems or free enzymes in solution in a stirred tank reactor. The system comprises of single reactor of any of the said reactor types or plurality of reactors of any of the said reactor types arranged in series or in parallel. In some aspects, the enzyme is immobilized onto the solid matrix by means of adsorption or gel entrapment or covalent cross linking or any other process known in art. In certain aspects, the residence time of the feed solution in the different reactors of the isomerization system may be uniform or different. In some aspects, the reaction in the isomerization system is carried out in a pH between 5.0 to 9.0 and conducted at a temperature between 35 – 90 °C by using a suitable isomerase enzyme. Such isomerase includes enzymes, but not limited to those involved in the conversion of glucose to fructose.
In another embodiment, section 20 may comprise of a refining system, wherein the ionic impurities and colour are removed from the aqueous mixture of sugars primarily composed of glucose and fructose obtained from any of the above mentioned sources or a combination of different sources, comprising of decolorization columns containing decolorization resins, cation exchange columns containing cation exchange resins, anion exchange columns containing anion exchange resins and mixed bed columns containing cation and anion exchange resins. In some aspects, the operational sequence of columns in the refining system 1 can be altered to provide an aqueous mixture of sugars primarily composed of glucose and fructose having a conductivity of less of than 20 µS/cm. In certain aspects, the production of an aqueous solution rich in monosaccharides, primarily glucose and fructose could also utilize sucrose solutions or clarified juice from sugarcane processing plants.
In some aspects of the present invention, as shown in section 20 of Figure 1, the process for the production of aqueous solution rich in monosaccharides, primarily glucose and fructose, utilizes high fructose syrups and/or high fructose corn syrups and/ or fructose containing syrups. In this aspect, the sugar solutions with various weight compositions of glucose and fructose could be advantageously fed directly to the refining system 1 to provide an aqueous mixture of sugars primarily composed of glucose and fructose having a conductivity of less of than 20 µS/cm.
As shown in Figure 1, section 30 the process for producing allulose from sucrose and/ or high fructose syrups and/or high fructose corn syrups and/ or fructose containing syrups and/ or glucose syrups and/ or polysaccharides like starch and cellulose may comprise of epimerization system and refining system 2.
In certain embodiments of the present invention, the epimerization system as shown in section 30 of Figure 4, utilizes the aqueous solution rich in monosaccharides, primarily glucose and fructose, from section 20 to convert the fructose in the said aqueous solution to D-allulose by using a suitable epimerase enzyme. In this aspect, the epimerization system may comprise of immobilized enzymes in packed bed reactor systems or free enzymes in solution in a stirred tank reactor. The system comprises of single reactor of any of the said reactor types or plurality of reactors of any of the said reactor types arranged in series or in parallel. In some aspects, the enzyme is immobilized onto the solid matrix by means of adsorption or gel entrapment or covalent cross linking or any other process known in art.
In certain embodiments, the residence time of the feed solution in the different reactors of the allulose conversion system may be uniform or different. In some aspects, the enzymatic conversion reaction to produce D-allulose is carried out at a pH between 5.0 to 10.0 and conducted at a temperature between 35 – 90 °C. In this aspect, the aqueous solution emerging from the epimerization system comprises of glucose, fructose and D-allulose produced from the aqueous solution rich in monosaccharides, primarily glucose and fructose, from section 20 to convert a portion of the fructose in the said aqueous solution to D-allulose by using a suitable epimerase enzyme. Such epimerases include enzymes, but not limited to those involved in the conversion of fructose to allulose. In some aspects, the portion of fructose, in the aqueous solution rich in monosaccharides, primarily glucose and fructose, converted is at-least 20%.
In another aspect as shown in Figure 1, section 30, the refining system 2, wherein the ionic impurities and colour are removed from the aqueous solution emerging from the epimerization system comprising of glucose, fructose and D-allulose, is comprised of decolorization columns containing decolorization resins, cation exchange columns containing cation exchange resins, anion exchange columns containing anion exchange resins and mixed bed columns containing cation and anion exchange resins. In some aspects, the operational sequence of columns in the refining system 2 can be altered to provide an aqueous mixture of sugars primarily composed of glucose and fructose having a conductivity of less of than 20 µS/cm.
In some aspects of the present invention, the ternary separation chromatography system as shown in section 40 of Figure 4 relates to a process and system for separating components of a mixture comprising of more than two components by contact between solid and liquid phases in variable length chromatography zones. In particular, the invention relates to a chromatography system with multiple columns suitable for separation of sugars from an aqueous mixture comprising of more than two sugars like glucose, fructose and D-allulose with variable length chromatography zones are carried out by asynchronous switching mode. In an aspect of the invention in this system, every cycle time is divided into sub-cycles as represented in Table 1. In each sub-cycle the zone lengths are not fixed but are varied over time. The zone lengths oscillate continually by one column, with the decrease of one zone offsetting the increase of the adjacent zone and vice versa. The mode of operation using six column assembly is shown in Figure 2 and table 1.
The Table 1 corresponds to first cycle with feed inlet to column 1. Next cycle starts at the end of sub-cycle 4 of cycle 1 with inlet/outlet positions change by one column length as D(5)/E(5)/F(2)/R(3).
The present invention provides advantages over conventional process and products. The present invention provides an efficient multi-column chromatography separation process for separation of ternary mixtures with variable length chromatography zones.
In some aspects of the present invention, the ternary separation chromatography system as shown in section 40 of Figure 1 4, relates to a process and system for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and D-allulose. In this aspect, the system comprises of a continuous chromatography system having a plurality of columns positioned adjacent to each other and fluidically connected in series forming a closed loop configuration, wherein each column in the system is defined with a feed pump, diluent pump, extract pump and raffinate pump. More specifically, said system comprises of a plurality of columns that contain solid bed material arranged in series and in a closed loop, whereby said loop comprises a number of fluid injection lines in each column or column section that are connected to at least one injection pump and a number of fluid draw-off lines of each column or column section that are connected to at least one draw off means, at least one valve on each line, whereby said loop defines at least three chromatographic zones, whereby each of this is determined by a fluid injection point and a fluid draw-off point, whereby the system is characterized in that it comprises means for controlling the variation in time of the length of the zones that are connected to said valve and that are suitable for shifting by one column or column section the positions of the injection points and draw-off points in an intermittent manner.
According to one embodiment of the present invention, the continuous ternary separation chromatography system as shown in section 40 of Figure 1, is configured to receive the feed solution (1) containing a ternary mixture into at least one of the chromatography columns (C1-C6) through its corresponding feed pump and wherein the feed solution contains a ternary mixture comprising of allulose, one or more isomers of allulose and optionally, one or more additional sugars like glucose.
In yet another embodiment of the present invention, the continuous ternary separation chromatography system as shown in section 40 of Figure 1 4, is configured to receive the diluent into at least one of the chromatography columns (C1-C6) through its corresponding diluent pump wherein the diluent used is preferably water.
In yet another embodiment of the present invention, the continuous ternary separation chromatography system as shown in section 40 of Figure 4, is configured to continously withdraw an extract fraction from at least one of the chromatography columns (C1-C6) by its corresponding extract pump, wherein the extract fraction comprises of an aquous solution rich in D-allulose or sucrose isomers.
In another embodiment of the present invention, the continuous ternary separation chromatography system as shown in section 40 of Figure 4, is configured to continously withdraw the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) by its corresponding raffinate pump, wherein the raffinate 1 fraction comprises of an aqueous solution rich in glucose and raffinate 2 comprises of a mixture rich in glucose and fructose with neligible allulose.
According to an embodiment of the present invention, the shifting of the positions of the injection points and draw-off points is in the same direction as that of the flow in the columns. In yet another embodiment of the present invention, the flow rate of the fluid that circulates in a given zone is generally constant. In another embodiment of the present invention, during the cycle time the position of the injection or draw-off points is shifted with non-constant time shift.
According to yet another embodiment the present invention provides a process, during the cycle time of which it is possible to shift all the injection and draw-off positions with a constant time and advantageously with a time phase shift that is at-least equal to a quarter cycle time.
According to another embodiment of the present invention, the means of draw off at the extract and raffinate lines is by using a pump. Alternatively, an analog valve connected to a flow meter can also be used.
In accordance with an aspect of the invention, the columns (C1-C6) in the continuous ternary separation chromatography system as shown in section 40 of Figure 4, are filled with the chromatographic bed material, wherein the bed material is an ion exchange resin. Examples of suitable chromatographic bed materials include but not limited to, resins that are strong acid cation resins. Some examples of suitable strong acid cation resins include product sold by Dow Chemicals Co (Dowex99 Ca/320; Dowex99 Ca/310 and Dowex99 Ca/280), Mitsubishi Chemical Co. (UBK 555) and Purolite (PCR642 Ca).
In certain embodiments, present invention relates to a process and system for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose wherein the continuous ternary separation chromatography system as shown in section 40 of Figure 4, is operated as per the scheme provided in Table 1 - VariCol configuration for 6 column arrangement corresponding to first cycle. The process comprises the steps of:
i. Sub-cycle 1: Providing the feed sugar mixture comprising of glucose, fructose and allulose to column 1 (C1) and simultaneously drawing off the raffinate from column 2 (C2) and simultaneously providing the eluent to column 4 (C4) and drawing-off the extract from column 4 (C4) resulting in zone I comprising of one column length, zones II and III comprising of two columns each and zone IV comprising of a single column;
ii. Sub-cycle 2: After a certain point in time (t+Td2) advantageously shifting the extract drawing-off point alone by one column from column 4 (C4) to column 5 (C5) while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are now comprised of two columns and zone II and zone IV are comprised of single column;
iii. Sub-cycle 3: After a certain point in time (t+Td3) advantageously shifting the eluent inlet to column 5 (C5) from column 4 (C4), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone II are now comprised of one column and zone III and zone IV are comprised of two columns;
iv. Sub-cycle 4: After a certain point in time (t+Td4) advantageously shifting the feed inlet from column 1 (C1) to column 2 (C2), while the remaining inlet and outlet points in the previous step remain unaltered resulting in changes of the zone lengths whereby zone I and zone III are comprised on one column and zone II and zone IV are comprised on two columns. The four steps correspond to cycle 1;
wherein after a certain point in time, initiating cycle 2 by one column increment at inlet and outlet valve positions and the sub-cycles 2, 3, and 4 follow the same pattern as in cycle 1; and
continuing the operation following the above scheme for an extended number of cycles for continuous separation of components.
According to another embodiment of the present invention, the number of columns or column sections is temporarily zero in one or more zones of the system.
In an embodiment of the present invention, for a given total column number, infinite possible configurations with the said process would depend on the phase shift of the switching that are carried out.
Yet another embodiment of the present invention, relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose wherein glucose, fructose and allulose are preferably in the range of 10 – 40%, 15 – 40% and 1 – 25% respectively.
In another embodiment, the present invention relates to a process for separation and purification of sugars from a sugar mixture comprising of glucose, fructose and allulose wherein cycle switching time provided is in between 50-120 min. According to another aspect of the embodiment of the present invention, the temperature of the system is maintained at a temperature ranging between 30°C – 90 °C.
The continuous ternary separation chromatography system as shown in section 40 of Figure 4, results in three streams, an aqueous stream rich in glucose, an aqueous stream rich in glucose and fructose with negligible D-allulose and an aqueous stream rich in D-allulose.
The aqueous streams rich in glucose and rich in glucose and fructose with negligible D-allulose are advantageously recycled back to section 20 as shown in Figure 4. In this aspect, the fraction rich in glucose is recycled to the isomerization step, wherein the glucose present in the solution is converted to aqueous mixture of sugars primarily composed of glucose and fructose. In this aspect, the isomerization system may comprise of immobilized enzymes in packed bed reactor systems or free enzymes in solution in a stirred tank reactor. The system comprises of single reactor of any of the said reactor types or plurality of reactors of any of the said reactor types arranged in series or in parallel. In some aspects, the enzyme is immobilized onto the solid matrix by means of adsorption or gel entrapment or covalent cross linking or any other process known in art. In certain aspects, the residence time of the feed solution in the different reactors of the isomerization system may be uniform or different. In some aspects, the reaction in the isomerization system is carried out in a pH between 5.0 to 9.0 and conducted at a temperature between 35– 90 °C by using a suitable isomerase enzyme. Such isomerase includes enzymes, but not limited to those involved the conversion of glucose to fructose.
In another aspect, the aqueous stream rich in glucose and fructose with negligible D-allulose obtained after the continuous ternary separation chromatography is recycled to section 20 and mixed with the glucose and fructose fraction obtained after the isomerization step. This fraction is directly used as the feed for the epimerization step after a concentration step. This operation mode avoids the requirement of an additional chromatographic separation process to generate fructose rich fraction for the epimerization step. The mixing of the aqueous stream rich in glucose and fructose with negligible D-allulose obtained after the continuous ternary separation chromatography system is recycled with the glucose and fructose fraction obtained after the isomerization step also increases the total sugar concentration to the concentration step and results in reduced energy for the concentration step.
The aqueous fraction rich in D-allulose is further sent to downstream processing section 50 as shown in Figure 1 4, to produce crystals of D-allulose.
As shown in section 50 of Figure 1 4, the downstream processing section of D-allulose is comprised of concentration systems, D-allulose refining systems, crystallization systems among others.
As shown in section 50 of Figure 1 4, the concentration system 2 is comprised of single effect or multiple effect evaporation or membrane-based water removal systems to concentrate the aqueous D-allulose rich fraction, from section 40, from 1-15% w/w to final concentration of not less than 40% w/w.
In another aspect as shown in section 50 of Figure 1 4, the D-allulose refining system 3, wherein the ionic impurities and colour are removed from the aqueous solution primarily composed of D-allulose, is comprised of decolorization columns containing decolorization resins, cation exchange columns containing cation exchange resins, anion exchange columns containing anion exchange resins and mixed bed columns containing cation and anion exchange resins. In some aspects, the operational sequence of columns in the refining system can be altered to provide an aqueous mixture rich in D-allulose having a conductivity of less of than 20 µS/cm.
In an additional aspect, as shown in section 50 of Figure 1 4 the concentration system is comprised of single effect or multiple effect evaporation or agitated thin film evaporation or membrane-based water removal systems to concentrate the aqueous D-allulose rich fraction, from D-allulose refining system, from a concentration of above 40% w/w to final concentration of not less than 80% w/w.
In an additional aspect, as shown in section 50 of Figure 1, the crystallization system is comprised of a single or multiple agitated tanks or crystallizers which are operated in parallel or series to produce D-allulose crystals. In some aspects, the crystallizers are operated in batch mode or continuous mode. In some aspects, the crystals produced are continuously separated from the mother liquor or separated in batches using solid liquid separation systems like filtration or centrifugation systems.
The mother liquor obtained after the separation of D-allulose crystals from section 50 is advantageously recycled back to the continous ternary separation chromatography system through the refining system 2 and/ or to the concentration system 2 as shown Figure 1 4 to increase the overall conversion efficiency of the raw material to crystal D-allulose.
In an another aspect, the steps downstream of concentration system 3 as shown in section 50 of Figure 1 4, could be avoided to produce concentrated allulose syrup.
While the present invention has been described in terms of its specific embodiments, certain modifications and equivalents will be apparent to those skilled in the art and are included within the scope of the invention. The examples included herein are provided to illustrate particular aspects of the disclosure and do not limit the scope of the present invention.

Example 1:
Separation of ternary mixture comprising of glucose, fructose and allulose using variable length chromatography zones
The chromatography system as shown in figure 3 was used for the experiment. The columns (C1-C6) were filled with chromatography resin. After which the columns were filled with water. Then the hot water circulation was initiated in the jackets of the column and preheaters to maintain a temperature of 50 – 60 °C in the columns.
The feed pump, diluent pump and recycle pumps were started and a flow rate of 170 – 180 ml/min, 540 – 560 ml/min and 20 – 28 l/hr were maintained respectively. After the stabilization of the system the extract and raffinate pumps were started with a flow of about 330 – 360 ml/min. Each Sub cycle time varied from 2-32 min. After attaining steady state the collection of extract and raffinate 1 and raffinate 2 were initiated. The extract and raffinates collected separately for six cycles were pooled and analyzed for sugar composition using HPLC. The data has been provided here for 114 cycles.
Details of feed used
Feed data
Cycle Purity of sugars, % Concentration, w/v%
Allulose Fructose Glucose Allulose Fructose Glucose Total
1-72 12.57 37.28 50.15 6.8 20.17 27.13 54.09
73-114 12.51 37.35 50.14 6.64 19.82 26.61 53.06

Results: Total mass balance
Feed Extract Raffinate 1 Raffinate 2
Allulose, kg 50.71 48.41 0.16 0.7
Fructose, kg 151.02 0.32 19.06 118.04
Glucose, kg 202.9 0 68.49 124.3
Recovery of Allulose in Extract, % 96.35
Recovery of Fructose in Raffinate-2, % 85.25
Recovery of Glucose in Raffinate-1, % 35.21
Avg. Purity of Allulose in extract % 99.34

Example 2
Separation of ternary mixture comprising of isomaltulose, trehalulose, sucrose, fructose and glucose using variable length chromatography zones
The chromatography system as shown in figure 3 was used for the experiment. The columns (C1-C6) were filled with chromatography resin. After which the columns were filled with water. Then the hot water circulation was initiated in the jackets of the column and preheaters to maintain a temperature of 50 – 60 °C in the columns.
The feed pump, diluent pump and recycle pumps were started and a flow rate of 60 – 70 ml/min, 140 – 150 ml/min and 10 – 15 l/hr were maintained respectively. After the stabilization of the system the extract and raffinate pumps were started with a flow of about 60 – 80 ml/min. Each Sub cycle time varied from 2-32 min. After attaining steady state the collection of extract and raffinate 1 and raffinate 2 were initiated. The extract and raffinates collected separately for six cycles were pooled and analyzed for sugar composition using HPLC. The data has been provided here for 101 cycles.
Feed data
Isomaltulose Trehalulose Fructose Glucose Sucrose
Concentration, w/v 39.15% 7.14% 5.06% 4.31% 0.65%
Purity% 69.5% 12.7% 9.0% 7.7% 1.2%

Results – Total mass balance
Feed Extract Raffinate 1 Raffinate 2
Isomaltulose, kg 200.28 194.92 5.37 0.00
Trehalulose, kg 36.52 2.09 34.07 0.37
Fructose, kg 25.90 0.00 1.39 24.50
Glucose, kg 22.05 0.00 5.51 16.54
Sucrose, kg 3.34 0.00 0.10 3.24
Recovery of isomaltulose in extract 97.32%
Purity of isomaltulose in extract 98.94%
Recovery of trehalulose in raffinate 1 93.29%
Recovery of other sugars in raffinate 2 86.34%

ADVANTAGES
Following are advantages of the process.
• Three component (ternary separation) separation has been achieved with variable length chromatography zones (varicol)
• Superior separation with allulose or isomaltulose purity of more than 98%
• High recovery of component (allulose or isomaltulose) in the extract fraction – more than 95%
• The process and system offer very high productivity leading to a reduction in the overall resin reduction of close to 50%.
• Reduces the need for multiple chromatography systems for separating the said sugar mixture.
,CLAIMS:1. A process for separating and purifying sugar component(s) from the ternary mixture using continuous ternary separation chromatography system with variable length chromatography zones comprising;
i. introducing a feed solution containing a ternary mixture selected from monosaccharides and/or disaccharides into at least one of the chromatography columns having a chromatographic bed material through its corresponding feed pump;
ii. receiving the diluent/eluent into at least one of the chromatography columns through its corresponding diluent pump;
iii. continuously withdrawing an extract fraction from the chromatography columns (C1-C6) through its corresponding extract pump;
iv. continuously withdrawing the raffinate 1 or raffinate 2 from at least one of the chromatography columns through its corresponding raffinate pump; and
wherein said chromatography system comprises plurality of columns positioned adjacent to each other and fluidically connected in series forming a closed loop configuration and configured to operate with variable length chromatography zones by asynchronous switching of its inlet and outlet ports.

2. The process as claimed in claim 1, wherein said chromatography system is configured to operate in sub-cycles comprising;
i. Sub-cycle 1 wherein at time (t) providing the ternary mixture to column 1 (C1) and simultaneously drawing off the raffinate from column 2 (C2) and simultaneously providing the diluent to column 4 (C4) and drawing-off the extract from column 4 (C4) resulting in zone I comprising of one column length, zones II and III comprising of two columns each and zone IV comprising of a single column;
ii. Sub-cycle 2 wherein after a certain point in time (t+Td2) shifting the extract drawing-off point alone by one column from column 4 (C4) to column 5 (C5) while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone III comprises of two columns and zone II and zone IV comprising of single column;
iii. Sub-cycle 3 wherein after a certain point in time (t+Td3) shifting the eluent inlet to column 5 (C5) from column 4 (C4), while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone II comprises of one column and zone III and zone IV comprising of two columns;
iv. Sub-cycle 4 wherein after a certain point in time (t+Td4) shifting the feed inlet from column 1 (C1) to column 2 (C2), while maintaining the remaining inlet and outlet points in the previous step unaltered resulting in changes of the zone lengths whereby zone I and zone III comprises one column and zone II and zone IV comprising two columns;
wherein after a certain point in time, initiating cycle 2 by one column increment at inlet and outlet valve positions and the sub-cycles 2, 3, and 4 follow the same pattern as in cycle 1; and
continuing the operation following the above scheme for an extended number of cycles, for continuous separation of components.

3. The process as claimed in claim 1, wherein said chromatography system comprises of a plurality of columns that contain solid bed material arranged in series and in a closed loop, whereby said loop comprises a number of fluid injection lines in each column or column section that are connected to at least one injection pump and a number of fluid draw-off lines of each column or column section that are connected to at least one draw off means, at least one valve on each line, whereby said loop defines at least three chromatographic zones, whereby each of this is determined by a fluid injection point and a fluid draw-off point, whereby the system is characterized in that it comprises means for controlling the variation in time of the length of the zones that are connected to said valve and that are suitable for shifting by one column or column section the positions of the injection points and draw-off points in an intermittent manner.

4. The process as claimed in claim 1, wherein the ternary mixture comprises of monosaccharides and/or disaccharides and/or oligosaccharides or mixtures thereof.

5. The process as claimed in claim 1, wherein the diluent is water.

6. The process as claimed in claim 1, wherein the chromatography bed is an ion exchange resin preferably strong acid cation resin.

7. The process as claimed in claim 1, wherein during the cycle time of the process the position of the injection or draw-off points may be shifted with non-constant time.

8. The process as claimed in claim 1, wherein during the cycle time of the process the position of the injection or draw-off points may be shifted with a constant time with a time phase shift that maybe equal to a quarter cycle time.

9. The process as claimed in claim 1, wherein the flow rate of the fluid that circulates in each zone of the column is maintained constant.

10. The process as claimed in claim 1, wherein the feed stream has a flow rate ranging from 10 cm/h to 60 cm/h, the eluent has a flow rate ranging from 60 cm/h to 140 cm/h, the extract has a flow rate ranging from 20 cm/h to 90 cm/h, the raffinate has a flow rate ranging from 30 cm/h to 110 cm/h.

11. The process as claimed in claim 1, wherein the temperature of the continuous ternary separation chromatography system is maintained at a temperature ranging between 30°C – 90 °C.

12. The process as claimed in claim 1, wherein the shifting of the positions of the injection points and draw-off points is in the same direction as that of the flow in the columns.

13. A process for production of sugar component(s) comprising;
i. Section 20 comprising systems to generate an aqueous solution of raw material selected from monosaccharides, disaccharides, polysaccharides, oligosaccharides or mixtures thereof;
ii. Section 30 comprising of epimerization and refining systems for converting a part of the raw material from section 20 to an aqueous solution of ternary mixture of isomerized and/ or epimerized sugars comprising of monosaccharides and/or disaccharides without ionic impurities;
iii. Section 40 comprising of a continuous ternary separation chromatography system as claimed in any one of the preceding claims 1 to 12 for separation and purification of desired product from the mixture of isomerized and/or epimerized sugars, wherein the mixture of unreacted sugars as stream 1 and stream 2 are recycled to section 20 and the desired product as stream 3 for further processing in section 50;
iv. Section 50 comprising of systems for the downstream process of the desired product to obtain in crystal form comprising the concentration systems, demineralization and decolorization systems, crystallization systems, solid-liquid separation systems, and drying systems among others; and recycling the mother liquor obtained after the crystallization step to section 30 to increase the overall conversion of the raw material for continuous separation and purification of the desired component.
14. The process for production of D-allulose as claimed in any one of the preceding claims 1 to 13 comprising;
i. generating an aqueous solution of monosaccharides selected from glucose and fructose by enzymatic hydrolysis or acid hydrolysis or isomerization of starch and its derivatives like amylose, amylopectin, maltodextrin, maltose, fructose and glucose, Cellulose derivatives which include pretreated biomass, amorphous cellulose, cellodextrin, cellobiose and glucose, Sucrose derivatives which include glucose and fructose or from high fructose syrup (HFS) or high fructose corn syrup(HFCS) followed by concentrating in section 20;
ii. epimerizing the aqueous solution rich in monosaccharaides to obtain a mixture comprising of glucose, fructose and D-allulose and refining to remove ionic impurities in section 30;
iii. separating the glucose, fructose and D-allulose in the ternary sugar mixture in section 40 comprising the continuous ternary separation chromatography system with variable length chromatography zones configured to operate in sub-cycles resulting in a first process stream comprising an aqueous solution rich in glucose; a second process stream comprising a mixture of glucose and fructose; and a third process stream comprising an aqueous solution rich in D-allulose;
iv. recycling the first process stream rich in glucose to the isomerization step in section 20 to obtain the mixture of glucose and fructose;
v. mixing the second process stream with glucose and fructose of step (iv);
vi. subjecting the D-allulose obtained in third process stream to downstream process comprising of concentration(s), demineralization, decolorization, crystallization(s), solid-liquid separation and drying;
vii. recycling a portion of the mother liquor obtained after crystallization of step (vi) to the step of converting a portion of the fructose to D-allulose;
viii. recycling a portion of the mother liquor obtained after the crystallization step (vi) to the continuous ternary separation chromatography system (to obtain D-allulose rich fraction.

15. The process as claimed in claim 14, wherein the process in said chromatographic system comprises;
i. introducing a feed solution containing a ternary mixture selected from monosaccharaides and/or disaccharides and/or oligosaccharides or mixtures thereof into at least one of the continuous ternary separation chromatography columns (C1 to C6) having a chromatographic bed material through its corresponding feed pump
ii. receiving the diluent/eluent into at least one of the chromatography columns (C1-C6) through its corresponding diluent pump;
iii. continuously withdrawing an extract fraction enriched in D-allulose from any one of the chromatography columns (C1-C6) through its corresponding extract pump, and
iv. continuously withdrawing the raffinate 1 fraction comprising of an aqueous solution rich in glucose and raffinate 2 comprising of a mixture of glucose and fructose from at least one of the chromatography columns (C1-C6) through its corresponding raffinate pump.

16. The process as claimed in claim 15, wherein the concentration of glucose, fructose and allulose contained in the ternary mixture are in the range of 10 – 40%, 15 – 40% and 1 – 25% respectively.

17. A process for separation and purification of sucrose isomer from a sugar mixture comprising of one or more sucrose isomers, glucose and its isomers and one or more additional sugars including sucrose using the continuous ternary separation chromatography system as claimed in any one of the preceding claims 1 to 13 wherein the chromatography system is configured to continously withdraw an extract fraction from at least one of the chromatography columns (C1-C6) by its corresponding extract pump , wherein the extract fraction comprise of an aqueous solution rich in first sucrose isomer preferably isomaltulose.

18. The process as claimed in claim 17, wherein chromatography system is configured to continously withdraw the raffinate 1 or raffinate 2 from at least one of the chromatography columns (C1-C6) by its corresponding raffinate pump, wherein the raffinate 1 fraction comprise of an aqueous solution rich in second sucrose isomer preferably trehalulose and raffinate 2 comprising of a mixture of glucose and fructose.

19. The process as claimed in claim 18, wherein the concentration of first sucrose isomer or isomaltulose, second sucrose isomer or trehalulose and other sugars including monosaccharides are in the range of 30 – 90%, 3 – 30% and 2 – 30% respectively.