Crystalline D-allulose syrup

Abstract

A method for producing D-allulose syrup exhibiting high crystallinity is provided. [Solution] A D-allulose syrup containing, in addition to D-allulose, a D-allulose dimer mass content of less than 1.5% expressed in terms of dry mass, and characterized by the steps of providing a D-allulose composition containing D-allulose dimers, nanofiltering the D-allulose composition to provide a retentate and a permeate, recovering the nanofiltration permeate, and concentrating the permeate.

Description

[Technical Field]

[0001] The present invention relates to a D-allulose syrup, one of whose advantageous properties is that it can be crystallized more easily than prior art syrups. Another subject of the present invention relates to the use of this D-allulose syrup for producing food or pharmaceutical products. Another subject of the present invention relates to a method for producing this D-allulose syrup. [Background technology]

[0002] D-allulose (or D-psicose) is a rare sugar with 70% of the sweetness of sucrose. Contrary to sucrose, D-allulose is not metabolized by humans and therefore does not cause weight gain. It has a very low calorie content (0.2 kcal / gram), which prevents the increase of body fat. Furthermore, research has shown that D-allulose is non-cariogenic or even anti-cariogenic. Therefore, due to these properties, it has recently attracted great interest from the food industry and pharmaceutical industry.

[0003] D-Allulose is generally obtained enzymatically by reacting an aqueous solution of D-fructose with D-psicose epimerase, as described, for example, in application WO 2015 / 032761 A1 in the name of the applicant. Regardless of the enzyme used, the reaction is not complete, and the amount of fructose converted into D-allulose after epimerization is less than 30%.

[0004] Therefore, if it is desired to obtain a composition with a higher mass content of D-allulose, expressed in dry mass, it is necessary to carry out a step of separating D-allulose in order to separate it from the other components present, in particular fructose.To carry out this separation, the composition obtained by the epimerization reaction is chromatographed, very commonly for example by simulated moving bed continuous chromatography, which allows the isolation of a fraction enriched in D-allulose.

[0005] Japanese Patent Application Laid-Open No. 2001-354690 describes a method for purifying a D-allulose composition starting from a mixture of fructose and D-allulose, which comprises a separation step consisting of successive chromatographic steps using a specific order of sampling of various products from the mixture.A D-allulose-rich fraction (the D-allulose richness of which can reach 98%) and a fructose-rich fraction are recovered.The recovery rate of the D-allulose-rich fraction is 96%.

[0006] At the end of the above separation step, a liquid composition rich in D-allulose is obtained. These liquid compositions, commonly referred to as syrups, are used in the manufacture of food or pharmaceutical products. For example, WO 2015 / 094342, also in the name of the present applicant, describes the manufacture of a solid food product containing D-allulose syrup containing 50% to 98% D-allulose and natural proteins. Currently, various companies have announced the sale of D-allulose mainly in the form of this syrup.

[0007] WO 2016 / 135458 describes, for example, a syrup containing at least 80% allulose based on its dry mass, and its stability over time is studied. No manufacturing protocol for this syrup is described. The composition of the syrup was analyzed by high performance liquid chromatography, which is a standard method for analyzing this type of syrup. The syrup described in this application has been shown to have low crystallinity.

[0008] However, in some applications, there may be advantages in using D-allulose syrup that can be more easily crystallized. This is particularly the case in the production of biscuits, where the use of crystalline syrup allows the production of chewier biscuits. Another example is the production of chewing pastes with short textures, where the use of crystalline syrup is necessary to achieve a short texture in the mouthfeel of these chewing pastes. A syrup that is more easily crystallized can also increase the hardness of chewing gum, improving the texture in the mouth when chewed, or increase the hardness of caramel with a long texture. Summary of the Invention [Problem to be solved by the invention]

[0009] Through extensive research, the applicant has discovered that certain impurities are formed during the production process of D-allulose syrup. These impurities, to the applicant's knowledge, have not been reported in the literature. By using specific gas chromatography techniques, the applicant has been able to identify them as D-allulose dimers. The applicant has also been able to demonstrate that, contrary to other impurities such as glucose or fructose, these dimers have a very significant anti-crystallization effect. However, because these D-allulose dimers are formed during the process, their presence in D-allulose syrup is systematic, even if all precautions are taken during production. This causes the syrup to be weakly crystalline, thereby limiting its use, particularly in the aforementioned applications.

[0010] In addition to this observation, the applicant has also endeavored to provide novel syrups that exhibit higher crystallinity than those of the prior art, and to this end has developed a specific method that allows for the removal of a large amount of impurities from these D-allulose syrups. [Means for solving the problem]

[0011] A subject of the present invention is therefore a D-allulose syrup which, in addition to D-allulose, contains a mass content of D-allulose dimers determined by gas chromatography (GC) of less than 1.5%.

[0012] This syrup has the advantage that it crystallizes more easily than prior art syrups with the same dry matter content and D-allulose content, which makes it particularly suitable for use in the production of biscuits or chewing pastes.

[0013] Another subject of the invention is a method for producing a syrup according to the invention, comprising the steps of: providing an aqueous D-allulose composition comprising D-allulose dimers; nanofiltering the D-allulose composition to provide a retentate and a permeate; recovering the nanofiltration permeate; concentrating the permeate to provide a D-allulose syrup. The present invention relates to a method comprising:

[0014] By using a nanofiltration step, a permeate that is substantially free or even free of D-allulose dimers can be recovered, thereby obtaining the syrup of the invention after the concentration step.

[0015] As mentioned above, the applicant has been able to recognize that during the production of D-allulose syrup, certain impurities are systematically formed during the process. These impurities have not been reported in the literature to date. This is explained by the fact that these impurities are not detected on the chromatogram using high performance liquid chromatography techniques conventionally used to measure the purity of D-allulose (see Figures 4 and 5). The applicant was able to detect their presence using gas chromatography techniques (see Figures 6 and 7).

[0016] Regarding the document WO 2015 / 094342, this document uses a syrup containing D-allulose. However, it does not disclose how to prepare this syrup. In fact, as shown in the rest of this description, the selection of conditions for preparation, especially the concentration step, essentially affects the amount of D-allulose dimer formed, which has not been reported before. Therefore, the above document does not describe the syrup of the present invention. The same applies to the document WO 2016 / 135458 already mentioned, or to the document WO 2015 / 032761, which does not even exemplify such a syrup.

[0017] Regarding the aforementioned document JP 2001-354690 A, it describes the separation of fructose and allulose, and the chromatographic separation of a composition containing these two components.However, it does not describe the preparation of D-allulose syrup.In fact, the selection of conditions for the preparation of this syrup, especially for its concentration step, will essentially affect the amount of D-allulose dimer, and therefore this document does not describe the syrup of the present invention.

[0018] The documents WO 2011 / 119004A2, WO 2016 / 064087, CN 104447888 A and CN 103333935 A describe the preparation of crystalline compositions containing D-allulose, but do not describe the preparation of D-allulose syrup. The present invention also relates to D-allulose syrup obtainable by the method of the present invention. [Brief explanation of the drawings]

[0019] [Figure 1] 1 shows a circuit for producing crystalline allulose syrup. [Figure 2] 1 shows a circuit for producing crystalline allulose syrup with a recirculation loop. [Figure 3]1 shows the permeation, or flux, curves associated with the nanofiltration step as a function of the volume concentration factor. [Figure 4] 1 shows an HPLC chromatogram of a D-allulose-rich composition sampled in the method of the present invention before nanofiltration. [Figure 5] 1 shows an HPLC chromatogram of the permeate sampled in the method of the present invention, i.e., after nanofiltration. [Figure 6] 1 shows a GC chromatogram in the area characteristic of dimers of a D-allulose-rich composition sampled in the method of the present invention before nanofiltration. [Figure 7] 1 shows a GC chromatogram in the region characteristic of dimers of the permeate sampled in the method of the present invention, i.e. after nanofiltration. [Figure 8] 1 shows the dry matter content of the supernatant of D-allulose syrups after 1 month of storage at 4° C. and 15° C. as a function of D-allulose dimer content. DETAILED DESCRIPTION OF THE INVENTION

[0020] The D-allulose syrup of the present invention is an aqueous solution containing a small amount of D-allulose dimer. The terms "aqueous composition" or "aqueous solution" generally refer to a composition or solution in which the solvent is essentially water. The term "D-allulose dimer" refers to a compound containing D-allulose condensed with at least a second monosaccharide, the same or different. These dimers are, for example, dimers of the D-allulose-D-allulose type.

[0021] As mentioned above, the fact that the amount of D-allulose dimer contained in the syrup is very low, i.e. its mass content expressed in dry mass is less than 1.5% according to the invention, makes it possible to obtain a syrup with a higher crystallinity.

[0022] As shown by Figures 4-7, these dimers are detectable by GC but not during HPLC analysis. For this reason, the masses of the various components, expressed as dry masses, are systematically determined by GC in this application. To determine the amount of each species in the composition, the sample is generally subjected to a processing step to convert the various species present into methoximated trimethylsilyl derivatives. The masses of each species are expressed in this application based on the total dry mass, unless otherwise noted.

[0023] The amounts of glucose, fructose and allulose can be determined in a gas chromatograph fitted with an injector heated to 300°C, a flame ionization detector (FID) heated to 300°C and a 40 meter DB1 capillary column with an inner diameter of 0.18 mm and a film thickness of 0.4 μm, the column temperature programmed as follows: from 200°C to 260°C at a rate of 3°C / min, then from 260°C to 300°C at 15°C / min, and maintained at 300°C for 5 minutes.

[0024] The term "amount of D-allulose dimer" is intended to mean the difference between the total amount of dimers in a sample determined by GC and the amount of well-known dimers that may be present, such as glucose-glucose dimers, such as maltose and isomaltose.However, the amount of these glucose-glucose dimers is generally very small or even non-existent.For example, the mass of glucose-glucose dimers in the syrup of the present invention is generally less than 0.2%, and often less than 0.1%.

[0025] The possible amount of glucose-glucose dimers can be determined under the same conditions as described above for glucose, fructose and D-allulose: Hydrolysis of glucose-glucose dimers in the sample; determining, in the same chromatograph and under the same conditions, the amount of total glucose, said total glucose including the initial glucose, called free, and the glucose obtained by hydrolysis of glucose-glucose dimers; Subtract the initial glucose amount in the sample from this amount of total glucose It can be calculated by:

[0026] The total amount of dimers, as a part thereof, can be determined in a gas chromatograph under the same conditions as those described above, except that the column used is a 30 meter DB1 capillary column with an internal diameter of 0.32 mm and a film thickness of 0.25 μm, and the column temperature is programmed as follows: from 200°C to 280°C at a rate of 5°C / min, then maintained at 280°C for 6 minutes, then from 280°C to 320°C at a rate of 5°C / min, and maintained at 320°C for 5 minutes.

[0027] This method is described in more detail in the Examples section.

[0028] In addition to D-allulose, the D-allulose syrup contains a mass content of D-allulose dimers of less than 1.5% as determined by gas chromatography (GC). The D-allulose syrup of the present invention has a mass content of D-allulose dimers in the range of 0.1% to 1.4%, advantageously in the range of 0.2% to 1.3%, and preferentially in the range of 0.3% to 1.2%. It is possible.

[0029] D-allulose syrup has higher crystallinity when its D-allulose content expressed by dry mass is 75% or more.It can have a D-allulose content expressed by dry mass of 80% or more, for example 85% or more, particularly 90% or more.The higher the D-allulose content expressed by dry mass, the higher the crystallinity of the syrup.

[0030] The D-allulose syrup advantageously comprises, based on its dry weight: ·75%~99% D-allulose; · 0% to 25% D-fructose; · 0% to 10% glucose; D-allulose dimer in the range of 0% to 1.5% (limit values not included), for example in the range of 0.1% to 1.4%, advantageously in the range of 0.2% to 1.3%, preferentially in the range of 0.3% to 1.2% may include:

[0031] D-allulose syrup can have a dry matter content of more than 50%, for example, in the range of 65% to 85%, particularly 70% to 83%, for example, in the range of 75% to 82%. The higher the dry matter content, the easier the syrup can crystallize. However, if the dry matter content is high, the viscosity of the syrup may increase, which may make it difficult to handle.

[0032] D-allulose syrup has traditionally been providing an aqueous composition comprising D-allulose; concentrating the aqueous composition to form a D-allulose syrup. The compound is obtained by a method comprising:

[0033] The syrup of the present invention can be prepared by the method described in detail below, which includes a nanofiltration step before the concentration step.

[0034] This nanofiltration step allows to limit the amount of D-allulose dimer in the syrup of the present invention.This nanofiltration step is carried out before the step of concentrating the D-allulose-rich composition.Therefore, this step can provide a D-allulose syrup with a lower D-allulose dimer content than that obtained by the same method without this nanofiltration step.

[0035] In the nanofiltration step, which is essential to the method of the present invention, when the D-allulose composition is subjected to nanofiltration, · Permeate low in D-allulose dimers; Furthermore, the retentate is enriched in D-allulose dimers. Two fractions are formed:

[0036] In Figure 1, which shows a circuit for producing a syrup of the present invention, stream 6 represents the permeate and stream 9 represents the retentate. For illustrative but non-limiting reasons, unless otherwise stated, the streams shown in the remainder of the description refer to the streams of this Figure 1 production circuit.

[0037] The nanofiltration permeate is an intermediate that allows for the production of this stock solution.

[0038] The terms "D-allulose dimer-poor" and "D-allulose dimer-rich" , which is clearly related to the content of D-allulose oligomers in the nanofiltered composition.

[0039] The nanofiltration permeate is an intermediate that allows for the production of the D-allulose syrup of the present invention.

[0040] A subject of the present invention is therefore a method for producing a syrup, comprising the steps of: providing an aqueous D-allulose composition comprising D-allulose dimers; nanofiltering the D-allulose composition to provide a retentate and a permeate; recovering the nanofiltration permeate; concentrating the permeate to provide the D-allulose syrup of the present invention. The present invention relates to a method comprising:

[0041] To carry out the nanofiltration step used in the present invention, the composition to be nanofiltered is passed through a nanofiltration membrane, which generally has a dry matter content of 5% to 15%.

[0042] The temperature of the nanofiltered composition may be in the range of 10-80°C, typically in the range of 15-50°C, and often about 20°C.

[0043] The selection of the membrane used for this separation is well known to those skilled in the art. The nanofiltration membrane may have a cut-off threshold of less than 300 Da, preferably in the range of 150-250 Da. Ideally, the membrane has a MgSO4 rejection rate of at least 98%. This may be, in particular, a Dairy DK or Duracon NF1 type membrane manufactured by GE®.

[0044] The pressure applied to the membrane can also vary within wide limits and may range from 1 to 50 bar, preferably from 5 to 40 bar and most preferentially from 15 to 35 bar.

[0045] This nanofiltration step may be accompanied by a diafiltration stage.

[0046] Preferably, the volume concentration factor (VCF) of the nanofiltration is in the range of 2 to 20. This volume concentration factor is easily adjusted by one skilled in the art.

[0047] This nanofiltration step can be carried out continuously.

[0048] At the end of this nanofiltration step, the collected permeate may contain 0% to 1.2%, for example 0.05% to 1.0%, in particular 0.1% to 0.5% D-allulose dimers based on its dry mass.

[0049] It goes without saying that the process according to the invention can include other steps, such as those found in the above-mentioned conventional processes and described in detail below. The process according to the invention can also include additional purification steps and even steps of diluting or concentrating intermediates in order to adjust the dry matter content and thus carry out the various steps of the process according to the invention under the best conditions. All of these steps can be carried out consecutively.

[0050] The syrup of the present invention generally has a dry matter content of 50% or more. To increase the dry matter content (the permeate has a dry matter content of less than 50%), a concentration step must be carried out, during which the D-allulose dimer content can be increased. Since the formation of D-allulose dimers also occurs during this concentration step, it is preferable to select conditions that allow the amount of these dimers formed to be limited. For example, during the concentration step The concentration step is generally carried out under reduced pressure, for example, at a pressure in the range of 5 to 100 mbar, preferably 20 to 70 mbar. This reduced pressure reduces the temperature required for evaporation and shortens the time for this concentration step. It can be carried out at a temperature in the range of 30 to 80°C, advantageously in the range of 34 to 70°C, and preferentially in the range of 37 to 50°C. This concentration step can be carried out in a single-stage evaporator or a multi-stage evaporator, for example, a two-stage evaporator. At the end of the concentration step, a D-allulose syrup of the present invention is obtained, which contains less than 1.5% D-allulose dimer. Typically, the syrup of the present invention contains 0% to 1.2%, for example, 0.1% to 1.2%, in particular 0.2% to 1.0% or 0.3% to 0.8% D-allulose dimer.

[0051] The method of the present invention also includes a step of providing an aqueous D-allulose composition containing D-allulose dimers. The mass content of various components of the syrup (particularly possible D-allulose, D-fructose, and glucose) is mainly determined by the respective contents of these components contained in the provided aqueous D-allulose composition. For example, if the provided aqueous D-allulose composition has a high D-allulose content, the permeate and the syrup obtained from this permeate also have a high D-allulose content.

[0052] Conventional methods for producing D-allulose compositions containing D-allulose dimers include: providing a D-fructose solution; epimerizing the solution to form a D-allulose composition comprising D-fructose and D-allulose; Optionally, a chromatography step to enrich the D-allulose composition for D-allulose; Optionally concentrating the D-allulose-enriched composition. Includes:

[0053] Thus, according to the method of the present invention, a step of chromatography of a composition comprising D-allulose and D-fructose can be carried out to provide a D-allulose composition, which in this case is advantageously obtained by epimerization of a D-fructose solution.

[0054] The D-allulose composition obtained after the chromatography step, which has a higher D-allulose content than the composition obtained at the end of the epimerization step, contains D-allulose dimers. In addition to this D-allulose composition, a D-fructose or "raffinate"-enriched composition is also formed during this chromatography step.

[0055] The composition of D-fructose provided for the epimerization step (stream 1) can be a D-fructose syrup or a glucose / D-fructose syrup obtainable by dissolving D-fructose crystals in water. Preferentially, this composition comprises a glucose / D-fructose syrup containing at least 90% D-fructose, preferentially at least 94% D-fructose, by dry weight.

[0056] In one scheme shown in FIG. 2, the D-fructose composition provided for the subsequent epimerization step is a mixture (Stream 1′) of this D-fructose syrup and at least one recycled fraction (Stream 10 or 12), which may be a raffinate (all of the raffinate or a portion), which recycled fraction may contain a higher amount of D-allulose.

[0057] The D-fructose composition subjected to the epimerization step is · 0% to 10% D-allulose; ·70%-100% D-fructose; · 0% to 10% glucose; 0% to 15% D-allulose dimer may include:

[0058] The epimerization step is carried out using the D-fructose composition provided above, optionally after adjusting the dry matter content. This step is generally carried out at a dry matter content in the range of 30% to 60%, often 45% to 55%. A D-psicose epimerase enzyme or a composition containing this enzyme is introduced into the composition. This enzyme-containing composition can be a lyophilized product of a host microorganism that synthesizes D-psicose epimerase, optionally Bacillus subtilis, particularly one of the microorganisms described in WO 2015 / 032761 A1. The pH is adjusted, for example, to a range of 5.5 to 8.5 depending on the enzyme used. The reaction can be carried out by heating to a temperature in the range of 40 to 70°C, often 45 to 60°C. The reaction can last for 0.1 to 100 hours, for example, 0.2 to 60 hours. This reaction can be carried out, for example, on an enzyme column, which is also advantageous for carrying out this step continuously.To operate continuously, it is also possible to carry out the reaction in succession using several reactors.To carry out this epimerization step, the teachings of WO 2015 / 032761 A1 can be used in particular.

[0059] At the end of this reaction, a composition containing D-fructose and D-allulose is formed, generally with a D-fructose / D-allulose weight ratio ranging from 85 / 15 to 55 / 45, and often with a D-fructose / D-allulose weight ratio ranging from 80 / 20 to 60 / 40. This ratio depends on the epimerization parameters used and very obviously on the amounts of D-allulose and D-fructose in the D-fructose composition provided to the epimerization step, which may be particularly high in the case of recycle.

[0060] At the end of this epimerization step, if necessary, a filtration step can be carried out to recover any cell debris present, especially when a lyophilized product of the host microorganism is used. This step can consist of a microfiltration step. The microfiltered composition corresponds to stream 3, and the cell debris is recovered in stream 8.

[0061] The method of the present invention may also include further purification steps. Typically, a step of desalting the composition containing D-fructose and D-allulose (stream 3) is performed before the chromatography step. This can be done by passing the composition through one or more cation exchange resins (e.g., Dowex 88-type cation exchange resins), anion exchange resins (e.g., Dowex 66-type anion exchange resins), and cationic-anionic mixtures. In Figure 3, this composition corresponds to stream 4. The resulting composition containing D-fructose and D-allulose is then desalted to a resistance of typically 100 kΩ cm. -1 This desalting step can also be preceded by a decolorizing step of the composition comprising D-fructose and D-allulose, for example by passing it through a column containing activated carbon.

[0062] The composition comprising D-fructose and D-allulose (stream 4) can then be subjected to a chromatography step to provide at least one composition enriched in D-allulose and one composition enriched in D-fructose. In one preferred mode, which will be explained in detail in the following description, the composition comprising D-fructose and D-allulose to which the chromatography step is performed is a mixture of the composition obtained from the epimerization step (stream 4) and at least one recycled fraction (stream 4'), and this recycled fraction is then subjected to a chromatography step to provide at least one composition enriched in D-allulose. The recycled fraction optionally contains a higher amount of D-allulose.

[0063] The composition in which the chromatography step is carried out comprises, based on its dry weight, · 22%-45%, generally 23%-37% D-allulose; · 45%-75%, generally 46%-70% D-fructose; · 0% to 10% glucose; 2% to 10% D-allulose dimer may include:

[0064] Any continuous type of chromatography can be used to perform this chromatography step, particularly simulated moving bed (SMB), modified simulated moving bed (ISMB), partitioned modified simulated moving bed (DISMB), sequential simulated moving bed (SSMB), or Mitsubishi Nippon Chromatography (NMCI) type chromatography. Water is generally used as the eluent. The chromatograph can be equipped with several columns in series, for example, 4 to 8 columns. These columns contain ion exchange resins, such as cation resins for exchanging calcium ions. The dry matter content of the composition containing D-fructose and D-allulose can range from 40% to 70%, and is generally about 50%. The temperature of the composition during chromatography is generally in the range of 40 to 80°C, preferably in the range of 55 to 65°C. This chromatography is continued for as long as necessary to achieve sufficient separation, and can last for several hours.

[0065] At the end of this step, a D-allulose-rich composition (stream 5) is obtained, which may contain at least 80% D-allulose, advantageously at least 90%, based on its dry matter content. This D-allulose-rich composition may have a dry matter content in the range of 5% to 15%. At the end of this step, a raffinate (stream 10) is also obtained, which generally contains at least 75% D-fructose, often at least 80%, based on its dry matter content. This raffinate generally has a dry matter content in the range of about 15% to 30%.

[0066] The process according to the invention can include recycling at least a portion of the nanofiltration retentate (Stream 9 in Figure 2) and / or raffinate (Stream 10 in Figure 2), which can increase the amount of D-allulose dimer in the various compositions prior to the nanofiltration step.

[0067] For example, the D-allulose-rich composition (stream 5) obtained at the end of the chromatography may contain, based on its dry weight: ·80%~98% D-allulose; · 0% to 20% D-fructose; · 0% to 10% glucose; 0.5% to 5% D-allulose dimer may include:

[0068] The raffinate (stream 10) is, for its part, based on its dry mass: · 1% to 10% D-allulose; ·70%-99% D-fructose; · 0% to 10% glucose; 1.3% to 20% D-allulose dimer may include:

[0069] The amounts of the various components of the D-allulose syrup of the present invention depend on the D-fructose solubilizer used. The parameters can be easily adjusted by those skilled in the art by selecting the composition of the solution, the parameters of the epimerization step, and even the parameters of the chromatography step. These selections can provide an aqueous D-allulose composition containing the D-allulose dimer used in the present invention, thereby determining the final D-allulose, D-fructose, and glucose contents in the syrup of the present invention. For example, specific components can be increased by adding a liquid or solid composition to the permeate that has a higher purity for these components than the permeate. For example, D-fructose crystals can be added to increase the D-fructose content of the syrup.

[0070] The syrup of the present invention can be advantageously used in the production of food or pharmaceutical products. This can be used in the well-known uses of allulose, generally as a sweetener. Among the uses that can advantageously utilize the higher crystallinity of the D-allulose syrup of the present invention, mention can be made of chewing gum in the form of sticks or dragees, caramels with a long texture, candies and chewing tablets, biscuits, cookies, muffins, cakes, gelatin-based confectionery, and chewing pastes, especially chewing pastes with a short texture.

[0071] The present invention will now be illustrated in the following Examples section, which should not be construed as limiting the invention. [Example]

[0072] Analysis method Gas chromatography The gas chromatograph used is a Varian 3800 type. - Split-splitless injectors (with or without dividers); - Flame ionization detector (FID); - a computer system for processing signals from the detector; - Automatic sampler (type 8400) can be attached.

[0073] The various amounts are determined by gas chromatography in the form of methoxylated trimethylsilyl derivatives and then quantified by an internal calibration method.

[0074] Measurement of D-allulose, D-fructose and glucose content The response factors used are 1.25 for D-allulose and D-fructose and 1.23 for glucose. No other monosaccharides are detected.

[0075] Sample preparation In a tared dish, weigh 100-300 mg of the sample to be tested and 10 ml of an internal standard solution consisting of 0.3 mg / ml methyl α-D-glucopyranoside in pyridine. Remove 0.5 ml from the tared dish, place in a 2 ml pot, and evaporate to dryness under a nitrogen stream. Add 20 mg of methoxylamine hydrochloride and 1 ml of pyridine. Cap and place in a Reacti-therm® incubation system at 70°C for 40 minutes. Add 0.5 ml of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA). Heat at 70°C for 30 minutes.

[0076] Chromatography conditions Column: DB1 capillary, 40 meters, 0.18 mm inner diameter, 0.4 μm film thickness, composed of 100% dimethylpolysiloxane, non-polar (J&W Scientific reference). Reference number: 121-1043 Column temperature: Programmed from 100°C to 260°C at a rate of 3°C / min, then 15°C / min to 300°C, and maintained at 300°C for 5 min. Injector temperature: 300℃ Detector temperature: 300°C (range 10 -12 ) Pressure: 40 psi (constant flow) Vector Gas: Helium Injection mode: Split (Split flow rate: 100 ml / min) Injection volume: 1.0 μl

[0077] D-allulose, D-fructose, and glucose were detected in this order. The unknown D-allulose has a retention time of 39.5 to 40 minutes under these conditions.

[0078] Measurement of D-allulose dimer and glucose-glucose dimer contents The response factors used are 1.15 for D-allulose dimers and maltose, and 1.08 for isomaltose. No other glucose dimers were detected.

[0079] Sample preparation: In a tared dish, weigh 100-300 mg of the sample to be tested and 10 ml of an internal standard solution consisting of 0.3 mg / ml phenyl β-D-glucopyranoside in pyridine.

[0080] Remove 0.5 ml from the tared dish, place in a 2 ml pot and evaporate to dryness under a stream of nitrogen.

[0081] 0.5 ml of a 40 g / l solution of hydroxylamine hydrochloride in pyridine is taken, stoppered, stirred and kept at 70° C. for 40 minutes.

[0082] Add 0.4 ml of BSTFA and 0.1 ml of N-trimethylsilylimidazole (TSIM). Heat at 70°C for 30 minutes.

[0083] Chromatography conditions Column: DB1 capillary 30 meters, 0.32 mm inner diameter, 0.25 μm film thickness (J&W Scientific reference number: 123-1032) Column temperature: programmed from 200°C to 280°C at a rate of 5°C / min (hold for 6 min), then 5°C / min to 320°C, hold at 320°C for 5 min. Injector temperature: 300℃ Detector temperature: 300°C (range 10 -12 ) Pressure: 14psi (constant flow) Vector Gas: Helium Injection mode: Split (Split flow rate: 80 ml / min) Injection volume: 1.2 μl

[0084] Viewing Results: The contents of the various components are expressed in g per 100 g of crude product and are calculated according to the following formula:

number

[0085] If the percentage value obtained for one of the components (expressed here on a crude basis) is greater than 20%, the sample is diluted and the GC analysis resumed in order to obtain a mass less than 20%.

[0086] The mass expressed on a crude basis is then expressed on a dry basis by dividing by the dry matter content of the sample being tested.

[0087] Since the characteristic peaks do not co-elute, the masses of D-allulose, D-fructose and glucose can be easily determined.

[0088] The maltose and D-allulose dimer peaks may co-elute, but it should be noted that maltose is never present in the syrups of the present invention and those described in the Examples below.

[0089] If no characteristic peak for maltose is detected, the surface area Si of the D-allulose dimer is determined by integrating the unknown peak between 10 and 17 minutes. If a characteristic peak for maltose is detected (as may be the case for the syrup of the present invention), the amount of maltose is determined and this amount is subtracted from the total amount of dimer.

[0090] To determine the total amount of glucose-glucose dimer, the samples are subjected to the following protocol. Hydrogen chloride hydrolysis Weigh approximately 50-500 mg of sample (adjust the weight depending on the expected sugar content) into a 15 ml hydrolysis tube with a Teflon screw cap, add 2 ml of internal standard solution (5 mg / ml galactitol in osmotic water) using a two-marked pipette, add 3 ml of water and 5 ml of 4N HCl solution.

[0091] The tube is sealed with a stopper and stirred on a vortex stirrer for 1 minute, and then placed in a thermostatic dry bath adjusted to 100°C for 1 hour, with occasional vortex stirring.

[0092] Desalination and concentration After cooling, the entire hydrolysate is placed in a 50 ml beaker. Add 6-8 g of a 50 / 50 mixture of 4 and AG50 W 8. Stir magnetically for 5 minutes. Filter through a paper filter. Collect the liquor and repeat this desalting step until a pH close to that of water is obtained.

[0093] Sample preparation In a tared dish, weigh 100-300 mg of the sample to be tested and 10 ml of an internal standard solution consisting of 0.3 mg / ml methyl α-D-glucopyranoside in pyridine. Remove 0.5 ml from the tared dish, place in a 2 ml pot, and evaporate to dryness under a nitrogen stream. Add 20 mg of methoxylamine hydrochloride and 1 ml of pyridine. Cap and place in a Reacti-therm® at 70°C for 40 minutes. Add 0.5 ml of BSTFA. Heat at 70°C for 30 minutes.

[0094] The total glucose content of the solution (including the initial glucose, called "free", and the glucose obtained by hydrolysis, particularly associated with the presence of lutose and isomaltose) is determined by GC analysis of glucose. The amount of maltose and the amount of D-allulose dimer, by subtracting it from the total amount of dimers assigned to the peak at 10-17 min, are then more easily estimated.

[0095] Example 1: Implementation of a continuous industrial production process for D-allulose syrup Example 1 comprises a continuous process for the production of crystalline D-allulose syrup. The process steps used are detailed in Figure 1. The compositions and flow rates of the streams in steps 1 to 5 are shown in Table 1a.

[0096] Step 1: 17.3 metric tons of Fructamyl D-fructose syrup (Tereos) (Stream 1) containing 95% D-fructose at 50% dry matter content (DM) was delivered to a 14m 3 Stream 1 is maintained at 55°C. Stream 2 is introduced into a stirred batch reactor having a working volume of 3.3 x 10 7 It is introduced into the tank in an amount sufficient to have a unit activity. Three reactors are used in series to continuously provide a syrup (Stream 2) consisting essentially of fructose and allulose at a flow rate of 360 kg / h.

[0097] The reaction conditions are as follows: ·Temperature: 55℃ pH=7 Response time: 48 hours

[0098] At the end of the reaction, the resulting stream 2 contains a richness of about 25% D-allulose and a richness of about 75% D-fructose.

[0099] Step 2: During batch operation, stream 2 is passed through a microfiltration membrane. Stream 3 is obtained, free of cell debris, along with the microfiltration retentate (stream 8), containing debris from the Bacillus subtilis lyophilisate, which is flushed out of the circuit. The microfiltration parameters are as follows: Transmembrane pressure: 0~3bar ·Pore diameter: 0.1μm ·Temperature: 50℃ ·Average flow rate: 15L / h / m 2 ·Membrane: Sepro PS35 Volume concentration factor: 33

[0100] Step 3: Desalting of stream 3 occurs over Dowex 88 strong cation resin followed by Dowex 66 weak anion resin at an average flow rate of 2 BV / h. The carboy is maintained at a temperature of 45°C, and the resistivity of stream 4 at the end of desalting is still 100 kΩ cm at the outlet (stream 4). -1 If this is not the case, regenerate the resin.

[0101] Step 4: Stream 4 is fed to a continuous chromatograph (SCC ARI® equipped with 8 columns) in a circuit. The average feed flow rate is 348 kg / h at 50% DM.

[0102] The chromatographic parameters are defined as follows: Volume / column: 2m 3 Resin: Dowex Monosphere 99Ca / 320 ·Temperature: 60℃ Flow rate: Water / Stream 4 (vol. / vol.): 2.4 Load: 0.09h -1

[0103] Two fractions are extracted: a raffinate (stream 10) and a D-allulose-rich fraction (stream 5) that proceeds to step 5. Stream 10 is bled off.

[0104] Step 5: Stream 5 is passed through a nanofiltration membrane in batch mode with the following parameters: Transmembrane pressure: 30 bar ·Temperature: 20℃ ·Membrane: GE Duracon NF1 8040C35 Volume concentration factor (VCF): 2.5

[0105] The allulose dimers are concentrated in the retentate (stream 9), which is flushed out of the circuit, while the permeate (stream 6) is collected. Figure 6 shows the details of the permeation of the syrup as a function of VCF.

[0106] Step 6: Stream 6 is passed through a two-stage evaporator, the internal pressure of which is less than 50 mbar. The first stage is at 38°C and allows the dry matter content to increase to 35%. In the second stage, a dry matter content of 77% is reached. At the end of this step, a D-allulose syrup (stream 7) is obtained.

[0107] The properties of the inventive syrup (Stream 7) are reproduced in Table 1b.

[0108] [Table 1]

[0109] [Table 2]

[0110] If the nanofiltration step is not carried out in the same process, the D-allulose syrup obtained is similar except that it contains 1.5% of D-allulose dimers (dialulose), expressed by dry mass.

[0111] Example 2: Evaluation of the crystallinity of various D-allulose syrups Example 2 involves the preparation of various D-allulose syrups with a dry matter content of 77% and a richness of D-allulose of about 95%.

[0112] These syrups are prepared by adjusting the volume concentration factor of nanofiltration step 5 to obtain different diallurose contents, and / or by preparing the syrup and mixing the resulting permeate or retentate with D-allulose crystals or D-fructose crystals, and / or by using nanofiltration membranes with a lower rejection threshold. This allows the content of various components to be adjusted while maintaining a D-allulose content of approximately 95%. The dry matter composition of the syrups can be seen in Table 2.

[0113] To evaluate the crystallinity of the syrups, sieved D-allulose crystallization initiator with an average particle size of 70 μm is introduced into each syrup at a rate of 0.3% (mass of initiator / mass of dry substance).

[0114] Each syrup thus initiated is then placed in a refrigerator at either 4°C or 15°C for 4 weeks.

[0115] At the end of this period, the dry matter of the supernatant (or mother liquor) of the sample is measured using the Karl Fischer method. The more crystallized the syrup, the lower the dry matter content of the supernatant (because the dry matter of the syrup is concentrated in D-allulose crystals).

[0116] [Table 3]

[0117] These results are shown in FIG.

[0118] These studies show that, contrary to, for example, glucose or fructose, the dimer of D-allulose is the compound that significantly affects the crystallinity of D-allulose syrups containing it.

[0119] Therefore, the lower the amount of D-allulose dimer in the syrup, the more crystalline the D-allulose syrup will be, even if the amount of D-allulose is still similar.

[0120] Example 3. Preparation of caramel with a long texture A caramel composition with a long texture (hard caramel) is prepared from the D-allulose syrup according to the present invention (sample 2) according to the formula in Table 3.

[0121] [Table 4]

[0122] Conclusion: It has been shown to be possible to obtain a caramel with a long texture while at the same time having the advantage of being low in calories.

[0123] Example 4. Preparation of chewing gum A chewing gum composition is prepared from the D-allulose syrup of the present invention (Sample 2) according to the following formulation.

[0124] Gum base (1) 28.0% Allulose powder 58.0% Allulose syrup 3.5% Mannitol powder (average diameter: 160μm) (2) 3.7% Mannitol powder (average diameter: 50 μm) (3) 0.1% Maltitol syrup (4) diluted to 60% dry matter 3.5% Strong Sweeteners (5) 0.2% Fragrance 3.0% (1) Optima (registered trademark), Cafosa (2) Pearlitol® 160C, Rockete (3) Pearlitol® 50 C, Rockete (4) Lycasin® 80 / 55, Rockete (5) GumSweet®, a sweetener solution

[0125] The gum base and allulose and mannitol powders are placed in a chamber at 55°C for 4 hours. The gum base, 60% of the allulose powder, mannitol powder and half of the maltitol syrup are mixed in a gum mixer at 50°C. The intense sweetener, the remaining allulose powder, the remaining maltitol syrup, allulose syrup and flavor are then added successively. The mixer is stopped and the resulting composition is removed and then formed into chewing gum sticks.

[0126] This chewing gum stick has the advantage of exhibiting a good texture when chewed, which is explained by its high hardness.

Claims

1. D-allulose syrup, the dry mass of which is determined by gas chromatography (GC) and contains a mass content of D-allulose of 75% or more, a mass content of D-allulose dimer of 0.1% to 1.2%, and a mass content of glucose of 0.1% to 10%, and has a dry matter content of more than 50%.

2. Based on its dry mass, - a mass content of D-allulose between 75% and 99%; - D-fructose with a mass content of between 0% and 24.8%; a mass content of glucose between 0.1% and 10%; - D-allulose dimer with a mass content ranging from 0.1% to 1.2% The D-allulose syrup according to claim 1, characterized in that it contains:

3. 3. D-allulose syrup according to claim 1 or 2, characterized in that it has a dry matter content ranging from 65% to 85%.

4. 3. D-allulose syrup according to claim 1 or 2, characterized in that it has a dry matter content ranging from 70% to 83%.

5. 3. D-allulose syrup according to claim 1 or 2, characterized in that it has a dry matter content ranging from 75% to 82%.

6. 6. The D-allulose syrup according to any one of claims 1 to 5, characterized in that it contains D-allulose dimers at a mass content ranging from 0.2% to 1.0%.

7. D-allulose syrup according to any one of claims 1 to 5, characterized in that it contains D-allulose dimers at a mass content ranging from 0.3% to 0.8%.

8. A method for producing the D-allulose syrup according to any one of claims 1 to 7. 、 - chromatographing the composition containing D-allulose and D-fructose obtained by epimerization of the D-fructose solution to obtain a D-allulose composition containing D-allulose dimers; nanofiltering the D-allulose composition to provide a retentate and a permeate; - recovering the permeate of said nanofiltration; - concentrating the permeate to provide a D-allulose syrup according to any one of claims 1 to 7 A method comprising:

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