Method for purifying isomaltulose from isomaltulose mother liquor

Abstract

The invention relates to a method for purifying isomaltulose from isomaltulose mother liquor. The method comprises the following steps: mixing an isomaltulose mother solution with isopinotriose hydrolase, carrying out a reaction, carrying out enzyme deactivation and solid-liquid separation after the reaction, collecting a clear liquid, and filtering to obtain a purified isomaltulose solution. According to the method, a brand new process is designed, the isomaltulose mother liquor is treated by utilizing the isopinotriose hydrolase, so that miscellaneous sugars such as isopinotriose and cane sugar in the mother liquor can be effectively decomposed to generate target products such as isomaltulose and monosaccharide, the separation difficulty is reduced, the miscellaneous sugars in the mother liquor can be effectively removed by combining a separation process, the concentration of the isomaltulose is increased, and the yield of the isomaltulose is increased. More isomaltulose is obtained from the mother liquor, meanwhile, a byproduct seaweed kelose is obtained, and the product yield and economic benefits are improved.

Description

Technical Field

[0001] The invention belongs to the technical field of separation and purification of functional sugars and relates to a method for purifying isomaltulose from isomaltulose mother liquor. Background Art

[0002] Isomaltulose, also known as Palatinose, is a natural disaccharide found in nature, such as sugar cane and honey. It is a reducing disaccharide formed by the α-1,6 glycosidic bond between glucose and fructose. Its sweetness is about 42% to 50% of sucrose. It has low glycemic index (GI), low insulin index, continuous energy supply, inhibition of fat accumulation, and no tooth decay. These characteristics of isomaltulose make it an ideal substitute for sucrose, and it has broad application prospects in the fields of food, chemical industry and medicine.

[0003] Isomaltulose is mainly produced by the conversion of sucrose isomerase. The conversion liquid is treated in a certain way and then crystallized and separated. Pure isomaltulose is extracted, and the remaining components will eventually remain in the separation mother liquor. When the impurities in the mother liquor reach a certain concentration, the isomaltulose product in the mother liquor will not be able to crystallize. In particular, the presence of miscellaneous sugars affects the quality of isomaltulose, and a large amount of isomaltulose remaining in the mother liquor cannot be fully utilized. In addition to a high concentration of isomaltulose, the mother liquor mainly contains by-products trehalulose, isomelizitose and monosaccharides, as well as incompletely converted sucrose, and can usually only be sold at a low price. Further separation and purification of the mother liquor will effectively improve the economic benefits. For example, CN104017028A discloses a method for purifying isomaltulose and trehalulose from isomaltulose mother liquor, comprising the following steps: (1) centrifuging the isomaltulose mother liquor and taking the supernatant to obtain a supernatant mother liquor; (2) taking the supernatant mother liquor, adding activated carbon, adsorbing and separating the solid from the liquid, collecting the supernatant to obtain a decolorized mother liquor; (3) diluting the decolorized mother liquor, separating it with a strong acidic cation exchange resin, eluting it with distilled water, collecting the eluate, sterilizing it with a filter membrane, and combining the eluates with the same components to obtain isomaltulose liquid and trehalulose liquid, respectively.

[0004] In summary, extracting more isomaltulose products from isomaltulose mother liquor and obtaining by-product trehalulose at the same time, improving product yield and economic benefits are of great significance to the field of isomaltulose production. Summary of the invention

[0005] In view of the deficiencies in the prior art and actual needs, the present invention provides a method for purifying isomaltulose from an isomaltulose mother liquor, which can efficiently extract more isomaltulose products from the crystallized isomaltulose mother liquor, and at the same time obtain the by-product trehalulose, thereby improving the product yield and economic benefits.

[0006] To achieve this object, the present invention adopts the following technical solutions:

[0007] In a first aspect, the present invention provides a method for purifying isomaltulose from an isomaltulose mother liquor, the method comprising:

[0008] The isomaltulose mother liquor is mixed with isomaltulose hydrolase to react, and after the reaction, the enzyme is inactivated and the solid-liquid is separated, and the clear liquid is collected and filtered to obtain a purified isomaltulose solution.

[0009] The present invention designs a new process, utilizes isomaltulose hydrolase to treat isomaltulose mother liquor, can effectively decompose isomaltulose and sucrose and other miscellaneous sugars in the mother liquor to generate target product isomaltulose and monosaccharides, reduces the difficulty of separation, and combines with the separation process, can effectively remove the miscellaneous sugars in the mother liquor, and improve the purity of isomaltulose.

[0010] It can be understood that in the present invention, isomaltulose mother liquor refers to the liquid part from which crystals are removed during the centrifugation process after isomaltulose crystallization in the production process of isomaltulose, which is usually difficult to crystallize again and its main components are isomaltulose and trehalulose.

[0011] Preferably, the amino acid sequence of the isomelizitose hydrolase includes the sequence shown in SEQ ID NO.1.

[0012] SEQ ID NO.1:

[0013] .

[0014] In the present invention, the specific hydrolase can be used to achieve efficient conversion of miscellaneous sugars, which is beneficial to improving the purity of isomaltulose.

[0015] Preferably, the ratio of the isomaltulose mother solution to isomelizitose hydrolase is 1kg:(0.1-1)g, for example, it can be 1kg:0.2g, 1kg:0.4g, 1kg:0.5g, 1kg:0.6g, 1kg:0.7g, 1kg:0.8g or 1kg:0.9g, etc.

[0016] Preferably, the reaction temperature is 18-60°C, for example, 19, 20, 21, 22, 23, 25, 30, 35, 40, 45, 50, 55, 56, 57, 58 or 59°C, and the reaction time is 0-72h, for example, 0.5, 1, 2, 3, 5, 10, 15, 20, 25, 30, 40, 50, 60, 65, 66, 67, 68, 69, 70 or 71h.

[0017] In the present invention, specific reaction conditions are designed for the conversion process of miscellaneous sugars and isomelizitose hydrolase in isomaltulose mother liquor, which can effectively improve the conversion efficiency.

[0018] Preferably, the enzyme inactivation method comprises heating treatment, the heating treatment temperature is 60-90°C, for example, it can be 61, 62, 66, 70, 75, 80, 85, 86, 87, 88 or 89°C, and the time is 10-30 minutes, including but not limited to 11, 12, 13, 14, 15, 16, 17, 20, 25, 26, 27, 28 or 29 minutes.

[0019] In the present invention, the solid-liquid separation can adopt the solid-liquid separation method commonly used in the art, such as filter cloth separation, centrifugal filtration, etc. Preferably, the solid-liquid separation method includes centrifugal treatment, and the rotation speed of the centrifugal treatment is 8000-10000rpm.

[0020] Preferably, the filtering membrane is a nanofiltration membrane.

[0021] Preferably, the molecular weight cut-off of the nanofiltration membrane is 200 to 300 Daltons.

[0022] In the present invention, a specific filtering method is designed for the mother liquor after the isomaltulose hydrolase conversion, which can effectively separate impurities in the mother liquor and further improve the purity of isomaltulose.

[0023] Optionally, the mass concentration of isomaltulose in the isomaltulose mother solution is 30% to 80%.

[0024] As a preferred technical solution, the method for purifying isomaltulose from isomaltulose mother liquor comprises the following steps:

[0025] (1) mixing isomaltulose mother liquid and isomaltulose hydrolase at a ratio of 1 kg:(0.1-1) g, and reacting at 18-60° C., wherein the amino acid sequence of the isomaltulose hydrolase includes the sequence shown in SEQ ID NO.1;

[0026] (2) after the reaction, the enzyme is inactivated at 60-90° C., and solid-liquid separation is performed by centrifugation at 8000-10000 rpm;

[0027] (3) Collect the clear liquid and filter it using a nanofiltration membrane with a molecular weight cutoff of 200 to 300 Daltons to obtain a purified isomaltulose solution.

[0028] Compared with the prior art, the present invention has at least the following beneficial effects:

[0029] The present invention designs a new process, utilizes isomaltulose hydrolase to treat isomaltulose mother liquor, can effectively decompose isomaltulose and sucrose and other miscellaneous sugars in the mother liquor to generate target product isomaltulose and monosaccharide, reduces the difficulty of separation, and is combined with the separation process, can effectively remove the miscellaneous sugars in the mother liquor, improve the purity of isomaltulose, realize obtaining more isomaltulose from the mother liquor, and obtain byproduct trehalulose at the same time, improve product yield and economic benefit. DETAILED DESCRIPTION

[0030] The technical solution of the present invention is further described below by specific implementation methods. However, the following examples are only simplified examples of the present invention and do not represent or limit the scope of protection of the present invention. The scope of protection of the present invention shall be subject to the claims.

[0031] If no specific techniques or conditions are specified in the examples, the techniques or conditions described in the literature in the field or the product instructions are used. If no manufacturer is specified for the reagents or instruments used, they are all conventional products that can be purchased through regular channels.

[0032] In a specific embodiment of the present invention, isomaltulose is purified by taking the isomaltulose mother liquor in the process of preparing isomaltulose by conventional biological method as an example, and the isomaltulose mother liquor is detected by high performance liquid chromatography, and its sugar component is the relative proportion of liquid chromatography purity, wherein the sugar component is shown in Table 1.

[0033] Table 1

[0034] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 77％ 6.2％ 5％ 4.4％ 4.6％ 2.8％

[0035] Example 1

[0036] This embodiment provides a method for purifying isomaltulose from isomaltulose mother liquor, the method comprising:

[0037] 0.5 g of isomezitose hydrolase (amino acid sequence SEQ ID NO. 1) was added to 1000 g of the mother liquor, and the reaction was stirred at 30° C. for 48 h. The reaction was terminated and samples were taken for analysis of the sugar components. The results are shown in Table 2. It can be seen that isomezitose and sucrose can be decomposed.

[0038] Table 2

[0039] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 80.3％ 6.2％ 0.3％ 7.1％ 5.9％ 0.2％

[0040] The enzyme was inactivated by heating at 80°C for 20 min, and solid-liquid separation was performed. The clear liquid was collected and separated by a nanofiltration membrane (molecular weight cutoff 200 Daltons) to obtain a separated and purified isomaltulose solution. The sugar components were analyzed. The results are shown in Table 3. It can be seen that the purity of isomaltulose can be significantly improved, and trehalulose as a by-product can be obtained.

[0041] Table 3

[0042] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 90.8％ 6.7％ 0.3％ 1.1％ 0.9％ 0.2％

[0043] Example 2

[0044] This embodiment provides a method for purifying isomaltulose from isomaltulose mother liquor, the method comprising:

[0045] 0.1 g of isomezitose hydrolase (amino acid sequence SEQ ID NO. 1) was added to 1000 g of the mother liquor, and the reaction was stirred at 40° C. for 72 h. The reaction was terminated and samples were taken for analysis of the sugar components. The results are shown in Table 4. It can be seen that isomezitose and sucrose can be decomposed.

[0046] Table 4

[0047] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 80.4％ 6.1％ 0.5％ 6.9％ 5.6％ 0.5％

[0048] The enzyme was inactivated by heating at 60°C for 30 min, and solid-liquid separation was performed. The clear liquid was collected and separated by a nanofiltration membrane (molecular weight cutoff 300 Daltons) to obtain a separated and purified isomaltulose solution. The sugar components were analyzed. The results are shown in Table 5. It can be seen that the purity of isomaltulose can be significantly improved, and trehalulose as a by-product can be obtained.

[0049] Table 5

[0050] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 90.9％ 6.8％ 0.3％ 1.0％ 0.9％ 0.1％

[0051] Example 3

[0052] This embodiment provides a method for purifying isomaltulose from isomaltulose mother liquor, the method comprising:

[0053] 1000 g of the mother liquor was added with 1 g of isomezitose hydrolase (amino acid sequence SEQ ID NO. 1), and the reaction was terminated after stirring at 60° C. for 18 h. Samples were taken for analysis of the sugar components. The results are shown in Table 6. It can be seen that isomezitose and sucrose can be decomposed.

[0054] Table 6

[0055] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 80.6％ 6.2％ 0.3％ 7.0％ 5.7％ 0.2％

[0056] The enzyme was inactivated by heating at 90°C for 10 min, and solid-liquid separation was performed. The clear liquid was collected and separated by a nanofiltration membrane (molecular weight cutoff 300 Daltons) to obtain a separated and purified isomaltulose solution. The sugar components were analyzed. The results are shown in Table 7. It can be seen that the purity of isomaltulose can be significantly improved, and trehalulose as a by-product can be obtained.

[0057] Table 7

[0058] Element Isomaltulose Trehalulose Isomezitose glucose fructose sucrose purity 91％ 6.9％ 0.3％ 0.9％ 0.8％ 0.1％

[0059] Example 4

[0060] This example provides a method for purifying isomaltulose from isomaltulose mother liquor. Compared with Example 1, the only difference is that the enzyme conversion reaction temperature is 18°C.

[0061] The purity of the isomaltulose in the separated and purified isomaltulose solution is 88%.

[0062] Example 5

[0063] This example provides a method for purifying isomaltulose from isomaltulose mother liquor. Compared with Example 1, the only difference is that the enzyme conversion reaction temperature is 15°C.

[0064] The purity of the isomaltulose in the separated and purified isomaltulose solution is 84%.

[0065] Example 6

[0066] This example provides a method for purifying isomaltulose from isomaltulose mother liquor. Compared with Example 1, the only difference is that the enzyme conversion reaction temperature is 65°C.

[0067] The purity of the isomaltulose in the separated and purified isomaltulose solution is 85%.

[0068] Example 7

[0069] This example provides a method for purifying isomaltulose from isomaltulose mother liquor. Compared with Example 1, the only difference is that the molecular weight cut-off of the nanofiltration membrane is 100 Daltons.

[0070] The purity of isomaltulose in the separated and purified isomaltulose solution is 83%.

[0071] In summary, a new process is designed in the present invention, and isomaltulose mother liquor is treated with isomaltulose hydrolase, which can effectively decompose isomaltulose and sucrose and other miscellaneous sugars in the mother liquor to generate target product isomaltulose and monosaccharides, thereby reducing the difficulty of separation. Combined with the separation process, the miscellaneous sugars in the mother liquor can be effectively removed, the concentration of isomaltulose can be increased, and more isomaltulose can be obtained from the mother liquor, while the by-product trehalulose can be obtained, thereby improving the product yield and economic benefits.

[0072] The applicant declares that the above is only a specific implementation mode of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily thought of by those skilled in the art within the technical scope disclosed by the present invention shall fall within the protection scope and disclosure scope of the present invention.

Claims

1. A method for purifying isomaltulose from isomaltulose mother liquor, characterized in that: The method comprises: The isomaltulose mother liquor is mixed with isomaltulose hydrolase to react, and after the reaction, the enzyme is inactivated and the solid-liquid is separated, and the clear liquid is collected and filtered to obtain a purified isomaltulose solution.

2. The method for purifying isomaltulose from isomaltulose mother liquor according to claim 1, characterized in that: The amino acid sequence of the isomelizitose hydrolase includes the sequence shown in SEQ ID NO.

1.

3. The method for purifying isomaltulose from isomaltulose mother liquor according to claim 1 or 2, characterized in that: The ratio of the isomaltulose mother solution to the isomaltulose hydrolase is 1kg:(0.1-1)g.

4. The method for purifying isomaltulose from isomaltulose mother liquor according to any one of claims 1 to 3, characterized in that: The reaction temperature is 18-60° C. and the reaction time is 0-72 hours.

5. The method for purifying isomaltulose from isomaltulose mother liquor according to any one of claims 1 to 4, characterized in that: The enzyme inactivation method comprises heating treatment, wherein the temperature of the heating treatment is 60 to 90° C. and the time is 10 to 30 minutes.

6. The method for purifying isomaltulose from isomaltulose mother liquor according to any one of claims 1 to 5, characterized in that: The solid-liquid separation method includes centrifugal treatment, and the rotation speed of the centrifugal treatment is 8000-10000 rpm.

7. The method for purifying isomaltulose from isomaltulose mother liquor according to any one of claims 1 to 6, characterized in that: The filtering membrane is a nanofiltration membrane.

8. The method for purifying isomaltulose from isomaltulose mother liquor according to claim 7, characterized in that: The molecular weight cut-off of the nanofiltration membrane is 200 to 300 Daltons.

9. The method for purifying isomaltulose from isomaltulose mother liquor according to claims 1-7, characterized in that: The mass concentration of isomaltulose in the isomaltulose mother solution is 30% to 80%.

10. The method for purifying isomaltulose from isomaltulose mother liquor according to claims 1-9, characterized in that: The method for purifying isomaltulose from isomaltulose mother liquor comprises the following steps: (1) mixing isomaltulose mother liquid and isomaltulose hydrolase at a ratio of 1 kg:(0.1-1) g, and reacting at 18-60° C., wherein the amino acid sequence of the isomaltulose hydrolase includes the sequence shown in SEQ ID NO.1; (2) after the reaction, the enzyme is inactivated at 60-90° C., and solid-liquid separation is performed by centrifugation at 8000-10000 rpm; (3) Collect the clear liquid and filter it using a nanofiltration membrane with a molecular weight cutoff of 200 to 300 Daltons to obtain a purified isomaltulose solution.