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Title:
PROCESS FOR SEPARATING KETOSES
Document Type and Number:
WIPO Patent Application WO/2018/116272
Kind Code:
A1
Abstract:
The present disclosure relates to a process for separating D-Allulose from an aqueous feed mixture comprising D-Allulose and at least one other ketose, particularly, fructose. The process comprises: adding calcium chloride to the solution comprising D-Allulose and fructose to get a mixture; drying the mixture obtained in the above step; adding aqueous ethanol to the dried mixture to precipitate the fructose as a hydrated addition compound of said calcium chloride; filtering the precipitate formed and recovering the fructose from the precipitate; collecting the filtrate comprising calcium addition D-Allulose and recovering the D-Allulose from the filtrate.

Inventors:
RAY SIBNATH (IN)
SIL ANINDYA (IN)
PANDEY BANIBRATA (IN)
Application Number:
PCT/IB2017/058352
Publication Date:
June 28, 2018
Filing Date:
December 22, 2017
Export Citation:
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Assignee:
PETIVA PRIVATE LTD (IN)
International Classes:
C07H3/02; C13K11/00; C13K13/00
Foreign References:
US4692514A1987-09-08
CA1292988C1991-12-10
FI84839C1992-01-27
US4785794A1988-11-22
US3533839A1970-10-13
Attorney, Agent or Firm:
BHATTACHARYYA, Goutam et al. (IN)
Download PDF:
Claims:
The Claim:

1. A process for separating D-Allulose from a solution comprising a mixture of pscicose and fructose; wherein the process comprises the steps of:

adding calcium chloride to the solution comprising D-Allulose and fructose to get a mixture;

drying the mixture obtained;

adding aqueous ethanol to the dried mixture to precipitate the fructose as a hydrated addition compound of said calcium chloride;

filtering the precipitate formed and recovering the fructose from the precipitate;

collecting the filtrate comprising calcium addition D-Allulose and recovering the D-Allulose from the filtrate.

2. The process as claimed in claim 1, wherein the calcium chloride is added in an amount of from about 10% to about 100% by weight based on the total amount of the D- Allulose and fructose present.

3. The process as claimed in claim 1, wherein the calcium chloride is added in an amount of about 50% by weight based on the total amount of the D-Allulose and fructose present.

4. The process as claimed in claim 1, wherein the ratio of fructose to calcium chloride is from about 1:0.5 to about 1: 1.

5. The process as claimed in claim 1, wherein the drying is a spray drying.

6. The process as claimed in claim 5, wherein the spray drying is carried out in a spray drier at a pumping speed of about 5 RPM to about 200 RPM, in a vacuum of about -100 mmWC to about -200 mmWC, and at an aspiration speed of about 1350 RPM to about 2000 RPM.

7. The process as claimed in claim 6, wherein the inlet and outlet temperature of the spray drier is from about 100 °C to 140 °C and 40 °C to 70 °C, respectively.

8. The process as claimed in claim 1, wherein moisture content of the dried mixture is from about 0% to about 30%.

9. The process as claimed in claim 1, wherein the water content in the aqueous ethanol is from about 0.01 % to about 30%.

10. The process as claimed in claim 9, wherein the water content in the aqueous ethanol is about 5%.

11. The process as claimed in claim 1, wherein the recovery of fructose from the precipitate comprises the steps of:

treating the precipitate with a solution of sodium sulphate to get insoluble salt;

filtering the insoluble salt by filtration;

subjecting filtrate to ion exchange resin treatment; and

concentrating the resultant filtrate to 75-95% w/w sugar syrup; and crystallizing fructose.

12. The process as claimed in claim 11, wherein the crystallization process comprises:

dissolving said sugar syrup in ethanol at a temperature of about 50-70°C; seeding the solution with fructose; and

allowing crystallization to occur; and recovering crystalline fructose.

13. The process as claimed in claim 12, wherein the temperature is allowed to about 15-25 °C, after seeding, at a decreasing rate of 1 °C per hour, to initiate the crystallization of the fructose.

14. The process as claimed in claim 1, wherein the recovery of D-Allulose from the filtrate comprises the steps of:

treating the filtrate with a solution of sodium sulphate to get precipitate; filtering the precipitate formed;

stripping the alcohol from the filtrate;

subjecting the stripped filtrate to ion exchange resin treatment;

concentrating the resultant solution to about 75-95% w/w sugar syrup; and crystallizing D-Allulose.

15. The process as claimed in claim 14, wherein the crystallization process comprises:

dissolving said sugar syrup in ethanol at a temperature of about 50-70°C; seeding the solution with D-Allulose; and

allowing crystallization to occur; and recovering crystalline D-Allulose.

16. The process as claimed in claim 15, wherein the temperature is decreased to about 15-25 °C, after seeding, at a decreasing rate of 1 °C per hour, to initiate the crystallization of the D-Allulose.

Description:
"PROCESS FOR SEPARATING KETOSES"

This application claims the benefit of Indian provisional application number, 201641044094, filed on December 23, 2016 which is hereby incorporated by reference in its entirety.

FIELD OF THE INVENTION

The present invention relates to a process for separating ketoses. More specifically, the invention relates to a process for separating D-Allulose from an aqueous feed mixture comprising D-Allulose and at least one other ketose.

BACKGROUND OF THE INVENTION

The known processes for production of D-Allulose (earlier known as psicose), on a commercial scale have one or more disadvantages due to absence of economical methods for separating D-Allulose and fructose. The widely-used processes are: 1) Removal of fructose by yeast fermentation; and 2) Simulated Moving Bed (SMB) method using phenol- formaldehyde resin with Ca 2+ .

In SMB operation, costs of the machineries are expensive whereas utilization of fructose by yeast is not economical as nearly 70-80% of the D-Allulose and D-fructose mixture containing D-fructose. For example, in fermentation method the fructose converted to glycerol, ethanol and acetic acids depending on the fermentation condition and side products obtained from fermentation have less commercial value than fructose. Similarly, separation of D-fructose and D-Allulose in SMB methods has several disadvantages and they are: 1) Fructose-D-Allulose mixture should be ion free; and/or feed concentration should be low for getting better separation resolution and hence the separated product was much diluted. In other words, in this process large amount of effluent would be produced; 3) It involves large capital cost investment; and 4) Further downstream process requires removing the carrier leached, soluble phenol-formaldehyde before concentration followed by crystallization. Moreover, phenol- formaldehyde is carcinogenic in nature.

Therefore, there is a need to develop an alternate process for separating D- Allulose from an aqueous feed mixture comprising D-Allulose and at least one other ketose which is devoid of one or more above concerns. SUMMARY OF THE INVENTION

Accordingly, the present invention provides a process for separating D-Allulose from an aqueous feed mixture comprising D-Allulose and at least one other ketose, particularly, fructose. The process comprises: adding calcium chloride to the solution comprising D-Allulose and fructose to get a mixture; drying the mixture obtained in the above step; adding aqueous ethanol to the dried mixture to precipitate the fructose as a hydrated addition compound of said calcium chloride; filtering the precipitate formed and recovering the fructose from the precipitate; collecting the filtrate comprising calcium addition D-Allulose and recovering the D-Allulose from the filtrate.

Before the methods of the present disclosure are described in greater detail, it is to be understood that the methods are not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the methods will be limited only by the appended claims.

Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the methods. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges and are also encompassed within the methods, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the methods.

Certain ranges are presented herein with numerical values being preceded by the term "about." The term "about" is used herein to provide literal support for the exact number that it precedes, as well as a number that is near to or approximately the number that the term precedes. In determining whether a number is near to or approximately a specifically recited number, the near or approximating unrecited number may be a number which, in the context in which it is presented, provides the substantial equivalent of the specifically recited number.

Unless defined 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 the methods belong. Although any methods similar or equivalent to those described herein can also be used in the practice or testing of the methods, representative illustrative methods and materials are now described.

All publications and patents cited in this specification are herein incorporated by reference as if each individual publication or patent were specifically and individually indicated to be incorporated by reference and are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited. The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present methods are not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed.

It is noted that, as used herein and in the appended claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as "solely," "only" and the like in connection with the recitation of claim elements, or use of a "negative" limitation.

It is appreciated that certain features of the methods, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the methods, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination. All combinations of the embodiments are specifically embraced by the present invention and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace operable processes and/or devices/systems/kits. In addition, all sub-combinations listed in the embodiments describing such variables are also specifically embraced by the present methods and are disclosed herein just as if each and every such sub-combination was individually and explicitly disclosed herein.

As will be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present methods. Any recited method can be carried out in the order of events recited or in any other order which is logically possible.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows chromatogram of bio-converted fructose D-Allulose mixture by DPEase enzyme;

FIG. 2 shows chromatogram of fructose-D-Allulose mixture after spray drying;

FIG. 3 shows chromatogram of calcium addition fructose dehydrate solid compound after 95% aqueous ethanol extraction;

FIG. 4 shows chromatogram of calcium addition D-Allulose compound in 95% aqueous ethanol extract;

FIG. 5 shows chromatogram of fructose after crystallization; and

FIG. 6 shows chromatogram of D-Allulose after crystallization.

DETAILED DESCRIPTION OF THE INVENTION

To the accomplishment of the foregoing and related objects the process of this invention comprises the features herein are fully described.

In an embodiment, the present disclosure provides a process for separating D- Allulose from an aqueous feed mixture comprising D-Allulose and at least one other ketose. The process comprises:

adding alkaline earth metal salt to the solution comprising D-Allulose and at least one other ketose;

drying the mixture obtained in the above step;

adding alcohol to the dried mixture to precipitate corresponding alkaline earth metal-ketose complex;

filtering the precipitate formed and recovering the ketose from the precipitate;

collecting the filtrate comprising calcium addition D-Allulose and recovering the D-Allulose from the filtrate.

The term "ketose" refers to monosaccharides containing one ketone group (=0) per molecule. Examples of ketoses include, but are not limited to, ketohexose (all six- carbon, ketone-containing sugars, including fructose), ketopentose (all five-carbon ketone containing sugars, including xylulose and ribulose), ketotetrose (all four-carbon, ketose containing sugars, including erythrulose), and ketotriose (all three-carbon ketose containing sugars, including dihydroxyacetone).

In certain embodiments, the alkaline-earth metal salt may be halide of alkaline- earth metal. In certain embodiments, the alkaline-earth metal salt is calcium chloride.

It is found that fructose and D-Allulose have similar solubility in anhydrous ethanol and the solubility of both sugar fructose and D- Allulose is enhanced in anhydrous ethanol in the presence of calcium chloride. When some small amount of water is added to fructose-anhydrous calcium chloride-anhydrous ethanol system, a large amount of white crystalline precipitate is formed at room temperature. The precipitate is not dissolved even after temperature is raised to 50 °C. However, D-Allulose-Ca complex is completely soluble in anhydrous ethanol at the same temperature and even at 0 °C. Analysis showed that solid part (precipitate) contains calcium salt of fructose in dehydrated form and the liquid part (filtrate) contains predominantly D-Allulose (85- 90%) and rest of other composition in liquid is fructose. Based on these facts, a new process for separating D-Allulose from fructose is developed.

In another embodiment, the present disclosure provides a process for separating D-Allulose from a solution comprising a mixture of pscicose and fructose. The process comprises the steps of:

adding calcium chloride to the solution comprising D-Allulose and fructose to get a mixture;

drying the mixture obtained in the above step;

adding aqueous ethanol to the dried mixture to precipitate the fructose as a hydrated addition compound of said calcium chloride;

filtering the precipitate formed and recovering the fructose from the precipitate;

collecting the filtrate comprising calcium addition D-Allulose and recovering the D-Allulose from the filtrate.

In the present process, chemically or enzymatically converted fructose and D- Allulose mixture in solution is mixed with different mole ratio of calcium chloride with respect to fructose present in mixture. The sugar and salt mixtures of different final concentrations (30-90 % w/v) are concentrated by rotary evaporation to obtain different molality mixture with respect to total solutes. In certain embodiments, the molality range may be from about 70 to about 90 molal with respect to total solute. Typically, the sugar concentration is from about is 40-50% w/v. In certain embodiments, the total sugar to calcium chloride mole ratio is from about 1:0.5 to 1:2. In a further embodiment, the total sugar to calcium chloride mole ratio is 1: 1.

After the addition of calcium chloride to the solution of fructose and D-Allulose, the resultant mixture may be filtered to remove any undesired and insoluble materials present in the mixture. Any method known in the art can be used for the filtration. In certain embodiments, the mixture is filtered through a micro filtration device.

After the filtration, the resulting solution may be dried to achieve desired moisture values. Any known method for drying the filtrate can be employed. In certain embodiments, the resulting solution is dried to get powder using spray drying technique in which a spray drier is used. The inlet and outlet temperature of the spray drier is maintained in the range of 100 °C to 200 °C and 40 °C to 90 °C, respectively. In certain embodiments, the inlet and outlet temperature of the spray drier is typically maintained in the range of 100 °C to 150 °C and 40 °C to 50 °C, respectively. In a further embodiment, the inlet and outlet temperature of the spray drier is about 130 °C and about 40 °C, respectively. The spray drying can be carried out with a feed rate of about 15 RPM, aspirator speed varying from about 1350 RPM to about 2000 RPM, and under vacuum varying from about -100 mmWC to about -200 mmWC. Other process conditions during the spray drying may be maintained as follows: feed concentration: 30-70%; and vacuum: pumping speed: from about 5 RPM to about 200 RPM. The moisture content of resultant dried powder is from about 0% to about 30%. In certain embodiments, it is from about 2% to about 5%. In a further embodiment, it is about 4%.

The dried powder is then extracted with ethanol containing 0-30% water. In certain embodiments, the dried powder is extracted with about 95 % ethanol water system (ethanol 95% + water 5%). The amount of 95% ethanol can be used is one part to five parts, preferably, two parts with respect to sugar-salt powder produced. The temperature employed for the extraction is from about 0° to 35° C, preferably 10-25° C. At elevated temperature fructose also enters solution rendering its separation from D-Allulose incomplete. The optimum extraction time of D-Allulose-calcium complex from the fructose-calcium complex can be done for 1-6 h. In certain embodiments, it is 2-3 h.

The precipitate (dehydrated calcium salt of fructose) thus precipitated is very stable, nonhygroscopic, and readily filterable. Further, it is not very much contaminated with D-Allulose.

The calcium chloride used in the process may be any commercial product. Although, commercial products of calcium chloride contain various impurities such as iron chloride, aluminum chloride, zinc chloride and sodium chloride, these impurities are harmless in the practice of the present process. Even when 3-5% of these impurities are admixed with calcium chloride purposed no obstacle is experienced.

The ethanol used in the process may be any commercial ethanol or recovered ethanol which contains no water or at most a few percent of water.

The dehydrated fructose-Ca salt (precipitate obtained) can be isolated using any filtration method known in the art. In certain embodiments, it is filtered using 10-11 micron filter under vacuum. The solid is washed with ethanol to get rid of the absorbed D-Allulose-Ca complex in solid, if any. The purity of the solid obtained is about 97% to about 99%. The ethanoic filtrate contains predominantly D-Allulose-Ca salt ranging from about 75-90%, fructose about 10-15% and glucose about 4-5%. The conversion of fructose to glucose is observed depending on the temperature of extraction and duration of extraction.

To recover the fructose from the addition compound (precipitate, dehydrated fructose-Ca salt) obtained as above, it is dissolved in equal parts of water and calcium content. The calcium is removed by treating with equimoles of sulfates such as sodium sulphate and ammonium sulfate, followed by filtration of said calcium sulfate. In certain embodiments, sodium sulfate is used for double decomposition of calcium sulfate. Then, the filtrate is concentrated to remove water and then it is made about 95% solution with respect to ethanol to remove the other salt from the solution. The remaining salt is removed by using ion exchange resin (cation and anion exchange resin). Finally, aqueous solution of fructose solution is concentrated to about 85-90 molal by vacuum evaporation and two parts of absolute ethanol is added with respect to the syrup weight and heated to about 50-70°C to dissolve the fructose. Then, fructose seed of about 1% is added to the solution and the temperature is decreased to about 15-25 °C, at a decreasing rate of 1 °C per hour with constant stirring. After that, finally crystallization is performed at a temperature of about 15-25°C with gentle agitation and recovered the fructose crystals. The crystallization yield obtained is about 80-85% of total sugar and purity of fructose obtained is about 99.7%.

In order to recover the D-Allulose from alcoholic solution, equimolar saturated sodium sulfate solution is added to alcoholic solution of calcium adduct D-Allulose compound with constant stirring at room temperature and incubated for 1-2 h for calcium sulfate and sodium chloride precipitation. The precipitates are separated by filtration. Ethanol is recovered by vacuum distillation. Then, the D-Allulose enriched syrup is diluted to equal amount of water and remaining salt is removed passing aqueous solution of D-Allulose through cation and anion exchange column simultaneously. The color is removed passing the active charcoal column. Finally, ion free solution is filtered through the micron filter to remove the contaminant particles and aqueous solution of D-Allulose solution is concentrated to 85-90 molal by vacuum evaporation and two parts of absolute ethanol is added with respect to syrup weight and heated to about 50-70°C to dissolve the fructose into alcohol. The D-Allulose seed upto 1% is added to solution and the temperature is reduced to about 15-25°C at a decreasing rate of 1 °C per hour with constant stirring. The crystallization is initiated at a temperature of about 15-25 °C with gentle agitation. The crystallization yield obtained is about 70-85% of total sugar and purity of the D-Allulose obtained is about 99.7%.

In the above description, calcium chloride is particularly referred to as the inorganic compound which is capable of information of an addition compound with fructose in the process of this invention. But other form of calcium salt like calcium bromide and iodide deposited with fructose in presence of ethanol.

On the other hand, it was also found that in aqueous ethanol, strontium chloride is the only compound which exhibits the same behavior as calcium chloride among the compounds of the other members of the same alkaline earth metal group as calcium, because no magnesium compounds and barium compounds proved to deposit the addition compound under similar conditions. When strontium chloride is employed in place of calcium chloride for the formation of the addition compound with fructose in the process of the invention, the removal of strontium chloride from the addition compound may be carried out in the same manner as that for calcium chloride as previously described.

The present process is very advantageous to separate D-Allulose from fructose with any distribution proportion in their mixture. This separation process can be applicable for any monosaccharide, di-saccharide and oligosaccharide preferably to their structural differences. Mainly this process is advantageous to separate any sugar mixture containing pyranose and furnanose form. Calcium complex with furanose sugar forms stable complex in their mono- or di-hydrated forms, which is less soluble in anhydrous ethanol. Whereas, calcium complex of pyranose sugars forms is highly soluble in anhydrous ethanol.

The ethanol used in the present process can be reused for extraction and crystallization process.

Adsorbed ion in ion exchange resin can be regenerated to the raw material and reused for this separation process and thus the less effluent generated in this current inventive process

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

The articles "a" and "an" are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

As used herein, the term "comprises" or "comprising" is generally used in the sense of include, that is to say permitting the presence of one or more features or components. As used herein, the term "feed mixture" refers to a mixture containing one or more extract components and one or more raffinate components to be separated by this process.

As used herein, the term "raffinate" refers to liquid phase which is recovered after separating the insoluble matrix or compound.

The present disclosure is further described with reference to the following examples, which are only illustrative in nature and should not be construed to limit the scope of the present disclosure in any manner.

EXAMPLES

Example 1

In this example solubility of fructose, D-Allulose and their mixture in anhydrous ethanol in the presence of anhydrous calcium chloride was tested. Fructose and D-Allulose and mixture thereof were treated with calcium chloride. Then, small quantity of water was added and stirred at room temperature for few hours for calcium sugar complex formation and precipitation. The precipitate was filtered. The sugar composition and yield were analyzed by HPLC. Precisely, to 2 gm of fructose, 2 gm of D-Allulose and lgm each of fructose and D-Allulose was added 0.5 ml of water to make 80 molal syrup concentrations separately in beaker. To this, 9.5 ml of absolute ethanol followed by 0.8 gm of anhydrous calcium chloride were added and incubated at room temperature for three hours with constant stirring. A solid precipitated was separated in case of fructose itself and fructose D-Allulose containing mixture. No solid is observed in D-Allulose solution. The solid was separated by filtration and analyzed by HPLC. The HPLC analysis data and yield are shown in Table 1

Table 1: Solubility and extraction of calcium addition fructose and D-Allulose compound

Syste Mat Wa Ca Eth Fruc Solid Composition Liquid in erial ter Ch anol tose composition g) (% (fus (ml) (yiel Fruc D- Glu Fruc D- Glu

) ed) d) tose Allu cose tose Allu cose

(%) lose (%) (%) lose (%) (%) (%)

Fructo 2 0.5 0.8 9.5 98 100 0 0 97.7 2.3 se

D- 2 0.5 0.8 9.5 0 0 0 0 4.3 95.7 0 Allulos

e

Fructo 1+1 0.5 0.8 9.5 96 99.8 0.2 0 13.8 85 1.2 se+D- Allulos

e

Example 2:

In this example stoichiometry of calcium chloride over total sugar and water in ethanol concentration was optimized for separation of fructose and D-Allulose from its mixture. For this purpose, 100 ml of 45.6% brix containing fructose-D-Allulose (80:20) mixture (total solid content 45.6 g or 0.25 moles) were mixed with different amounts of calcium chloride to make different calcium-sugar complexes. FIG. 1 shows the chromatogram of bio-converted fructose D-Allulose mixture as starting material. Anhydrous calcium chloride used was 11.25 gm and 16.87 gm to 22.5 gm based on the fructose content on fructose-D-Allulose mixture. Different fructose to calcium mole ratios (1 :0.5, 1 :0.75 and 1: 1) were used. Aqueous solution of the sugar and calcium chloride mixture were spray dried to make as free flowing powder. Spray drying condition was used for this purpose are as follows:

Feed RPM: 15, Inlet Temperature: 130°C, Outlet Temperature: 40°C, Aspirator RPM: 1350, Vacuum applied: -100 mmWC. In case of 1 : 1 mole ratio free flowing powder obtained. The calcium content and sugar compositions analyzed by ICP-OES and HPLC method.

To optimize the separation of fructose and D-Allulose from their calcium addition compound, different aqueous content ethanol used. In this experiment 10 gm of spray dried powder and two parts of 85 %, 90% and 95% aqueous ethanol incubated at room temperature for 3 hours with constant stirring. Finally, the precipitate was filtered and washed with absolute ethanol. The yield of solid powder and composition analyzed. Mixed sugar calcium chloride and extraction optimization data summarized in Table 2.

Table 2: Solubility and extraction of calcium addition fructose and D-Allulose compound from its mixture after spray drying

Example 3:

In this example 2 L of fructose-D-Allulose (80:20) mixture (brix of the sugar solution 45%) containing 900 gm of total mixed with 444 gm of anhydrous calcium chloride solution in such a way that fructoseiCaCh mole ratio is 1: 1. The sugar and salt solution was prepared by constant stirring at room temperature for an hour and filtered through micron filtered to remove the dust particle. The filtered solution fed in spray dryer with pump speed of 5 RPM, inlet and outlet temperature was set to 130 °C and 40 °C, respectively. Vacuum applied -100 mmWC with Aspiration RPM 1350. The free- flowing powder obtained in this process was 1184 gm with recovery yield of 88 %. The loss observed due to formation of HC1 gas in spray drying process, which is recoverable. It was observed that 12 % weight loss observed due to formation hydrochloric acid (HC1) gases in drying process. In powder, sugar composition analyzed to check the degradation of product. No degradation of product composition was observed. However, 2-3% fructose converted in to glucose was observed. Figure 2, shows the chromatogram of fructose-D-Allulose mixture after spray drying. The data (HPLC) showed that the mixture contains D-Allulose (20.47%), fructose (78.28%) and glucose (1.25%). Material analysis data of fructose-D-Allulose mixture along with calcium chloride after spray drying is depicted in table 3 below. Table 3: Material analysis data of fructose D-Allulose mixture along with calcium chloride after spray drying

Example 4:

Separation of calcium addition fructose compound from the spray dried powder:

The powder as prepared in example 3 with the sugar composition (D-Allulose 20.47%, fructose 78.28% and glucose 1.25%) was used for separation of calcium addition compound of fructose by 5% aqueous content ethanol extraction. In this example, 1 kg of spray dried powder extracted with 2 L of 95% aqueous ethanol with constant stirring by overhead stirrer setting RPM 250 at 25 °C for three hours. The resultant precipitate was filtered under vacuum and washed with 200 ml of anhydrous ethanol and dried under vacuum. The weight of the solid obtained was 788 gm and purity of fructose obtained was 98.5 % and D-Allulose content is 1.5%. The calcium and sugar composition were analyzed by ICP-OES and HPLC methods, respectively. Figure 3 shows the sugar composition of solid material obtained after aqueous ethanol extraction. The analysis results are depicted in Table 4 below.

Table 4: Material analysis data of calcium addition fructose dehydrate compound after aqueous ethanol (95 % ethanol content):

Material Weight (gm)

Total 788

Fructose 552.2

D-Allulose 8.4 Water 56

Calcium 61.5

Example 5:

Recovery of Fructose from calcium addition fructose compound and its crystallization:

The solid obtained in Example 4 was used for recovery of fructose from its calcium complex compound. In this example, calcium equimolar sodium sulfate (Na 2 S04) (148 gm) was dissolved in 1.2 L of water by setting overhead stirrer at a speed of 250 rpm at room temperature. To it, 500 gm of calcium-fructosate solid (as described in example 4) was added slowly with constant stirring for one hour. The precipitate thus obtained was filtered under vacuum and washed with 250 ml of water spraying on top. Filtrate containing fructose and sodium chloride passed sequentially through one set of cation exchange, followed by two sets of anion exchange and one set of charcoal column with each column dimension of (100 cmx 5 cm) with feed flow rate of 10 ml per minute at 40 °C. Then, pH and conductivity were adjusted to 6.5 and 10 μ8/αη, respectively using 100 gm of cation and anion exchange mixed resin and filtered through micron filtered to remove undesired particles.

Thereafter, the ion free aqueous fructose solution was concentrated to 85 % (w/w) under vacuum for crystallization. The weight of the syrup obtained was 418 gm. For crystallization, 1 L absolute ethanol was added and incubated at 60 °C for 2 hours until fructose syrup dissolved completely. The fructose seed up to 1 % of total sugar was added to the solution and the temperature was reduced to 16 °C slowly at a rate of 1°C per hour with constant stirring. Then, the crystallization was initiated at 16 °C with gentle agitation. The crystallization yield obtained was 85% of total sugar and purity of the fructose obtained was 99.7% (shown in Figure 5) Example 6:

Recovery of D-Allulose from calcium addition compound and its crystallization:

2.2 L of ethanoic filtrate obtained from Example 4 was processed for recovery of D- Allulose from its calcium addition compound. The total sugar content in this mixture was 156 gm. HPLC analysis (as shown in FIG. 5) showed that composition is having D- Allulose (85.8%), fructose (11.5%), and glucose (2.6%),

In this example, the 2.2 L of ethanoic solution was treated with saturated aqueous solution of sodium sulfate. The saturated solution was prepared by dissolving 98.2 gm Na 2 S0 4 in 700 ml water at room temperature. Removal calcium from D-Allulose containing sugar solution was done by slow addition of saturated solution of Na 2 S0 4 with constant stirring at a speed of 200 RPM. The solution was incubated at room temperature for one hour for complete decomposition of sodium sulfate to calcium sulfate (CaS0 4 ) and sodium chloride (NaCl).

Calcium sulfate and NaCl precipitate were filtered under vacuum and washed with 250 ml of absolute ethanol. Ethanol from the filtrate was stripped off by fractional distillation and aqueous solution containing sugar and residual NaCl were passed through one set of cation exchange, followed by two set anion exchange and one set of charcoal column with each column dimension of (100 x 2.5 cm) with feed flow rate of 10 ml per minute at 40 °C. Then, pH and conductivity were adjusted to 6.5 and 10 μ8/αη, respectively using 50 gm of cation and anion exchange mixed resin and filtered through micron filtered to remove particles.

Thereafter, the ion free aqueous D-Allulose rich solution was concentrated to 85 % (w/w) by vacuum evaporation for crystallization. The weight of the syrup obtained was 183 gm. For crystallization, 450 ml absolute ethanol was added to the 85 molal sugar solutions and incubated at 60°C for 2 hours at constant stirring until D-Allulose rich syrup dissolved completely. The D-Allulose seed up to 1% of total sugar's weight was added to the solution and the temperature was slowly reduced to 16 °C, at a decreasing rate of 1°C per hour with constant stirring. Then, the crystallization was initiated at 16 °C with gentle agitation for 16 hours. The crystallization yield obtained was 80 % of total sugar and purity of the D-Allulose obtained was 97.1% and fructose was 2.1% (shown in Figure 6). The remaining liquid mixed with main stream of crystallization. Advantages of The Present Process:

1) The process is more efficient, simple and scalable

2) The reagents used are non-toxic and easily recoverable

3) The process produces less effluent

4) Calcium-fructose (1:2) and calcium-D-Allulose (1: 1) complex produced by the present process can be used as food supplement for osteoporosis and calcium deficiency

Although the foregoing invention has been described in some detail by way of illustration and example for purposes of clarity of understanding, it is readily apparent to those of ordinary skill in the art in light of the teachings of this invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended claims.

Accordingly, the preceding merely illustrates the principles of the invention. It will be appreciated that those skilled in the art will be able to devise various arrangements which, although not explicitly described or shown herein, embody the principles of the invention and are included within its spirit and scope. Furthermore, all examples and conditional language recited herein are principally intended to aid the reader in understanding the principles of the invention and the concepts contributed by the inventors to furthering the art, and are to be construed as being without limitation to such specifically recited examples and conditions. Moreover, all statements herein reciting principles, aspects, and embodiments of the invention as well as specific examples thereof, are intended to encompass both structural and functional equivalents thereof. Additionally, it is intended that such equivalents include both currently known equivalents and equivalents developed in the future, i.e., any elements developed that perform the same function, regardless of structure. The scope of the present invention, therefore, is not intended to be limited to the exemplary embodiments shown and described herein. Rather, the scope and spirit of present invention is embodied by the appended claims.