A process for the separation and purification of mannose from a mixture of sugars

Abstract

The application discloses a process for separating and purifying mannose from mixed sugar, comprising the following steps: mixing mixed sugar containing mannose and glucose with an ethanol aqueous solution with a concentration of 95 wt% or above, heating to dissolve the mannose, and separating to obtain glucose. The application utilizes the different solubilities of monosaccharides in ethanol at different temperature conditions to realize the separation of mixed high-value monosaccharides. Compared with the method for separating monosaccharides by using resins and column chromatography in other industrial production, the application is more green and environmental-friendly, and the separation cost is greatly saved. Moreover, ethanol is the only edible alcohol, so the application is safer and more reliable.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of separation and utilization of biomass-based mixed high-value sugar, and particularly relates to a process for separating and purifying mannose from mixed sugar. BACKGROUND

[0002] Mannose is an isomer of glucose and galactose. As a simple monosaccharide, D-mannose exists in a variety of forms. A large amount of D-mannose exists in plant and cell wall oligosaccharides, peaches, apples, and orange peels. Mannose not only binds to human glycoproteins to regulate the human immune system and glycoproteins, but also eliminates inflammation. In addition, mannose can be used as a screening agent for gene-transformed cells and a clinical nutritional agent. In daily life, mannose can exist as a fruit and vegetable preservative or be applied to health products and beverages as a sweet additive due to its unique solubility and anti-melting properties. Mannose is widely used in commercial ice cream production. Adding mannose to livestock feed can inhibit the infection of salmonella typhimurium in poultry. In summary, mannose has a wide range of application scenarios.

[0003] Currently, mannose is mainly extracted from natural products such as konjac, and the extraction cost is high. The price of mannose is as high as 200,000 yuan / ton. Chemical catalytic synthesis of mannose uses glucose as a substrate and a catalyst to isomerize and synthesize mannose. This method has the advantages of low cost and high production efficiency. However, chemical catalytic synthesis of mannose cannot completely convert glucose into mannose. Therefore, the reaction solution contains mixed sugar including glucose, fructose, and mannose. How to efficiently separate high-purity mannose is a key problem for further reducing production costs and improving the purity of mannose. Biological separation method usually uses bacteria to consume one type of sugar and retain another type of sugar to achieve separation. Although this process can effectively separate glucose and other monosaccharides, the cost is relatively high. The common industrial method for separating various monosaccharides is column chromatography and exchange resin method. For example, Chinese invention CN103992361A discloses a method for separating mannose and glucose by sequential simulated moving chromatography. In this method, mannose syrup prepared by glucose epimerization is adjusted to a concentration of 60% as raw material. The sequential simulated chromatography separation device includes 6 chromatography columns and expensive adsorbent resin. The equipment is complex and the eluent required for separation is expensive. However, column chromatography and exchange resin method are relatively expensive and complex to operate. At present, since mannose is extracted from natural plants, there is no mature technology for efficiently separating mannose and other monosaccharides in industry. Therefore, it is urgent to explore a simple, efficient, and low-cost method for separating mannose without introducing other substances. SUMMARY

[0004] The present application aims to solve the technical problems of the prior art and provide a process for separating and purifying mannose and / or glucose from mixed sugar.

[0005] To solve the above technical problems, the present application discloses the following technical solutions:

[0006] A method for separating and purifying mannose and / or glucose from mixed sugar, comprising the following steps:

[0007] (1) mixing the mixed sugar containing mannose and glucose with an aqueous ethanol solution with a concentration of 95wt% or more, heating to dissolve the mannose, adding glucose seed crystals first and then centrifuging after standing to room temperature to obtain glucose;

[0008] (2) concentrating the solution after separating the glucose, adding mannose seed crystals first and then standing to separate to obtain mannose.

[0009] The mixed sugar in the present application is a dry product obtained by catalytic hydrolysis of sugar alcohol industrial sources, which includes mannose and glucose in some embodiments, and also includes fructose in some embodiments; in some embodiments, the mass ratio of the mannose to glucose is 1:0.72-1.22, preferably 1:0.82-1.12, preferably 1:0.92-1.02, preferably 1:0.97; in some embodiments, the mass ratio of the mannose to fructose is 1:0.07-1.31, preferably 1:0.08-1.26, preferably 1:0.09-1.21, preferably 1:0.10-1.16, preferably 1:0.11.

[0010] In step (1),

[0011] The concentration of the aqueous ethanol solution is 96wt% or more, preferably 96%-98%, preferably 96%-97%, such as 96.5%.

[0012] The mass ratio of the mixed sugar to the aqueous ethanol solution with a concentration of 90wt% or more is 1:8.7-15.4, preferably 1:9.7-14.4, preferably 1:10.7-13.4.

[0013] The heating temperature is 40-60℃, preferably 45-55℃.

[0014] The heating time is 10-20min.

[0015] The heating is ultrasonic heating.

[0016] The amount of glucose seed crystals added is 2-5mg per 0.36g of glucose contained in the mixed sugar.

[0017] The separation after standing to room temperature is standing to room temperature for 2-4h, until a large amount of white solid, mostly glucose, is separated.

[0018] The centrifugation is 10000-12000r / min for 5-10min, and the white solid obtained by centrifugation is dried at 45-55℃.

[0019] In step (2),

[0020] The concentration is that the solution of separated glucose is subjected to rotary evaporation at a temperature of 50-60℃ and a rotation speed of 40-60r / min for 5-10min, and a trace of liquid is reserved.

[0021] The adding amount of the mannose seed crystal is 2-5mg per 0.35g of mannose contained in the mixed sugar.

[0022] The standing separation is that after standing at 0-30℃ for 2-10h, a large amount of white solid, mostly mannose, is separated, preferably standing at 15-25℃ for 2-10h, and preferably standing at 18-25℃ for 4-8h for solid-liquid separation.

[0023] According to the method provided by the application, the yield of the obtained glucose is 95% or more, the purity of the obtained glucose is 85% or more, the yield of the obtained mannose is 95% or more, and the purity of the obtained mannose is 85% or more.

[0024] Advantages: Compared with the prior art, the application has the following advantages:

[0025] (1) The separation of glucose and mannose in the application is separated by using high-concentration ethanol aqueous solution as a solvent, which is more green and environmentally friendly than the separation of monosaccharides by using resin separation and column chromatography in other industrial production, can be recycled, and the separation method is simple and easy to operate, greatly saving the separation cost.

[0026] (2) The mixed sugar is freeze-dried by the mixed solvent of ethanol and water in the application, which helps to protect the structure of the separated sugar at low temperature and reduces the content of sugar crystallization water, and is simple and easy to operate. BRIEF DESCRIPTION OF DRAWINGS

[0027] The above and / or other aspects of the application will become more apparent by describing in detail the application with reference to the attached drawings and specific embodiments.

[0028] Figure 1 The purity of the mannose obtained by standing separation of 0.36g of glucose, 0.35g of mannose and 0.04g of fructose at different temperatures.

[0029] Figure 2 The purity of the glucose sample obtained by separation of 0.36g of glucose, 0.35g of mannose and 0.04g of fructose at different ethanol amounts.

[0030] Figure 3 H1-NMR of glucose before and after separation.

[0031] Figure 4 This is the H1-NMR of mannose before and after separation. DETAILED DESCRIPTION

[0032] The experimental methods described in the following examples are conventional methods unless otherwise specified; the reagents and materials are commercially available unless otherwise specified.

[0033] The freeze-dried mixed sugar described in the following examples is a catalytic hydrolyzate of an industrial source of sugar alcohol. The contents of the three sugars in the hydrolyzate were measured, and the contents of glucose, mannose, and fructose in the hydrolyzate were 0.36 g, 0.35 g, and 0.04 g, respectively. The mixture was freeze-dried to obtain a freeze-dried mixed sugar.

[0034] The detection method of glucose and mannose after separation in the following examples is as follows:

[0035] Place 2-5 mg of the separated and dried glucose in a test tube, add 1-2 ml of water to dissolve it completely, dilute it 1000-5000 times, filter it, and take 200 μL of the solution for detection in an ion chromatograph.

[0036] Place 2-5 mg of the separated and dried glucose in a test tube, add 1-2 ml of water to dissolve it completely, dilute it 1000-5000 times, filter it, and take 200 μL of the solution for detection in an ion chromatograph.

[0037] Example 1

[0038] 1. Add 0.75 g of freeze-dried mixed sugar to 8 g, 9 g, and 10 g of 96.5% ethanol aqueous solution, respectively, and place the mixed sugar solution in an ultrasonic cleaner at 50°C for 20 min;

[0039] 2. Add 3 mg of glucose crystals as seed crystals and let it stand at room temperature for 3 hours until a large amount of white solid precipitates, mostly glucose;

[0040] 3. Centrifuge the solid-liquid mixture at 12,000 rpm for 5 minutes. Dry the resulting white solid at 55°C, weigh it, and perform a purity test.

[0041] 4. The separated solution was placed in a rotary evaporator at a temperature of 50°C and a speed of 50 r / min for 7 minutes, retaining a trace amount of liquid.

[0042] 5. Add 3 mg of mannose crystals, and stand at 4°C, 10°C, 20°C, and 30°C respectively for 6 hours until a large amount of white solid, mostly mannose, is precipitated. After solid-liquid separation, dry the precipitated solid at 55°C, weigh it, and test its purity.

[0043] The results are as follows:

[0044] 1. The purity of mannose separated at different temperatures in this example is shown in Table 1. Figure 1

[0045] After separating glucose crystals from 10 g of 96.5% ethanol aqueous solution, concentrate the liquid by rotary evaporation, and stand the precipitated mannose crystals at 4°C, 10°C, 20°C, and 30°C respectively.

[0046] As can be seen from Table 1 and Figure 1 the purity of mannose crystals is the highest at 20°C, which is 78.96%, indicating that 20°C is the most suitable temperature for the precipitation of mannose crystals.

[0047] Table 1 Separation results of mixed sugars at different crystallization temperatures with 8 g of ethanol solution

[0048]

[0049]

[0050] 2. The purity of glucose separated at different ethanol dosages in this example is shown in Table 2. Figure 2

[0051] After separating glucose crystals from 8 g, 9 g, and 10 g of 96.5% ethanol aqueous solution respectively, concentrate the liquid by rotary evaporation, and stand the precipitated mannose crystals at 20°C. Analyze the purity of glucose crystals and mannose crystals.

[0052] As can be seen from Table 2 and Figure 2 the lower the solution used, the higher the purity of glucose, but the quality of separated glucose crystals decreases more; the more solution used, the higher the purity of mannose, but the quality of separated mannose decreases, and the quality of mannose obtained using 8 g of 96.5% ethanol aqueous solution is 0.05 g less than that obtained using 9 g of 96.5% ethanol aqueous solution.

[0053] Table 2 Separation results of mixed sugars at different ethanol dosages with crystallization temperature at 20°C

[0054]

[0055] The H1-NMR results of glucose before and after separation in this example are shown in Table 3. Figure 3 ​​As shown, the peak at 6.1 and 6.5 ppm is characteristic of glucose, and the characteristic peak of glucose exists before and after separation, but the peak height is increased, which is probably due to the presence of bound water.

[0056] The H1-NMR results of mannose before and after separation in this example are shown in Figure 3. Figure 4 As shown, the peak at 6.2 ppm is characteristic of mannose, and the characteristic peak of mannose exists before and after separation, but the peak height is decreased, which is probably due to the presence of bound water. The content of fructose is less, and most of the fructose exists in the glucose and mannose sample after separation, so it is not detected.

[0057] Example 2

[0058] 1. Add 0.75 g of freeze-dried mixed sugar to 8 g of 96.5% ethanol aqueous solution, and place the mixed sugar solution in an ultrasonic cleaner at 50°C for 20 min;

[0059] 2. Add 3 mg of glucose crystals as seed crystals, and stand at room temperature for 3 h until a large amount of white solid, mostly glucose, is precipitated;

[0060] 3. Centrifuge the above solid-liquid mixture at 12000 r / min for 5 min, and weigh the white solid obtained after drying at 55°C for purity detection;

[0061] 4. The separated solution is placed in a rotary evaporator at a temperature of 50°C and a speed of 50 r / min for 5-10 min to retain a small amount of liquid;

[0062] 5. Add 3 mg of mannose crystals, and stand at 20°C for 6 h until a large amount of white solid, mostly mannose, is precipitated. After solid-liquid separation, the precipitated solid is dried at 55°C and weighed for purity detection.

[0063] Example 3

[0064] 1. Add 0.75 g of freeze-dried mixed sugar to 9 g of 96.5% ethanol aqueous solution, and place the mixed sugar solution in an ultrasonic cleaner at 50°C for 20 min;

[0065] 2. Add 3 mg of glucose crystals as seed crystals, and stand at room temperature for 3 h until a large amount of white solid, mostly glucose, is precipitated;

[0066] 3. Centrifuge the above solid-liquid mixture at 12000 r / min for 5 min, and weigh the white solid obtained after drying at 55°C for purity detection;

[0067] 4. The separated solution is placed in a rotary evaporator at 50°C and 50 r / min for 5-10 min, and a trace amount of liquid is reserved;

[0068] 5. 3 mg of mannose crystals are added, and the solution is left to stand at 20°C for 6 h until a large amount of white solid, mostly mannose, is precipitated. After solid-liquid separation, the precipitated solid is dried at 55°C and weighed, and the purity is tested.

[0069] Example 4

[0070] 1. 0.75 g of the freeze-dried mixed sugar is added to 9 g of 96.5% ethanol aqueous solution, and the mixed sugar solution is placed in an ultrasonic cleaner at 50°C for 20 min;

[0071] 2. 3 mg of glucose crystals are added as crystal seeds, and the solution is left to stand at room temperature for 3 h until a large amount of white solid, mostly glucose, is precipitated;

[0072] 3. The above solid-liquid mixture is centrifuged at 12000 r / min for 5 min, and the obtained white solid is dried at 55°C and weighed, and the purity is tested;

[0073] 4. The separated solution is placed in a rotary evaporator at 50°C and 50 r / min for 5-10 min, and a trace amount of liquid is reserved;

[0074] 5. 3 mg of mannose crystals are added, and the solution is left to stand at 10°C for 6 h until a large amount of white solid, mostly mannose, is precipitated. After solid-liquid separation, the precipitated solid is dried at 55°C and weighed, and the purity is tested.

[0075] Example 5

[0076] 1. 0.75 g of the freeze-dried mixed sugar is added to 9 g of 96.5% ethanol aqueous solution, and the mixed sugar solution is placed in an ultrasonic cleaner at 50°C for 20 min;

[0077] 2. 3 mg of glucose crystals are added as crystal seeds, and the solution is left to stand at room temperature for 3 h until a large amount of white solid, mostly glucose, is precipitated;

[0078] 3. The above solid-liquid mixture is centrifuged at 12000 r / min for 5 min, and the obtained white solid is dried at 55°C and weighed, and the purity is tested;

[0079] 4. The separated solution is placed in a rotary evaporator at 50°C and 50 r / min for 5-10 min, and a trace amount of liquid is reserved;

[0080] 5. Add 3 mg of mannose crystal, respectively, stand at 30 °C for 6 h, until a large amount of white solid is precipitated, mostly mannose, after solid-liquid separation, the precipitated solid is dried at 55 °C, weighed and the purity is tested.

[0081] The above-described embodiments only express several embodiments of the present application, which are described in more detail and in more detail, but cannot be understood as limiting the scope of the patent of the present application. It should be noted that for ordinary skilled in the art, without departing from the concept of the present application, a number of modifications and improvements can be made, which belong to the protection scope of the present application. Therefore, the protection scope of the patent of the present application should be subject to the appended claims.

Claims

1. A method for separating and purifying mannose and / or glucose from a mixed sugar, characterized in that: The following steps are involved: (1) mixing a mixed sugar containing mannose and glucose with an ethanol aqueous solution having a concentration of 95 wt % or more, wherein the mass ratio of mannose to glucose in the mixed sugar is 1:0.72-1.22, and the mass ratio of the mixed sugar to the ethanol aqueous solution having a concentration of 95 wt % or more is 1:8.7-15.4, dissolving the mannose by ultrasonic heating at 40-60° C., adding glucose seed crystals, and then standing at room temperature for 2-4 hours, and then centrifuging to obtain glucose, wherein the amount of the glucose seed crystals added is 2-5 mg / 0.36 g glucose; (2) The separated glucose solution is concentrated, mannose seed crystals are first added, and then the solution is allowed to stand at 0-30° C. for 2-10 hours to separate mannose; the amount of the mannose seed crystals added is 2-5 mg / 0.35 g mannose.

2. The method according to claim 1, characterized in that In step (1), the concentration of the ethanol aqueous solution is above 96 wt %.

3. The method according to claim 1, characterized in that In step (1), the concentration of the ethanol aqueous solution is 96% to 98%.

4. The method according to claim 1, wherein In step (1), the concentration of the ethanol aqueous solution is 96% to 97%.

5. The method according to claim 1, wherein In step (1), the mass ratio of the mixed sugar to the ethanol aqueous solution with a concentration of 95 wt % or more is 1:9.7-14.

4.

6. The method according to claim 1, characterized in that In step (1), the mass ratio of the mixed sugar to the ethanol aqueous solution with a concentration of 95 wt % or more is 1:10.7-13.

4.

7. The method according to claim 1, characterized in that In step (1), the heating temperature is 45-55°C; The heating time is 10 to 20 minutes.

8. The method according to claim 1, characterized in that In step (1), the separation is performed by standing at room temperature for 2 to 4 hours and then centrifuging at 10,000 to 12,000 r / min for 5 to 10 minutes.

9. The method according to claim 1, characterized in that In step (2), the separation is carried out by standing at 15 to 25° C. for 2 to 10 hours.

10. The method according to claim 1, characterized in that In step (2), the separation is solid-liquid separation after standing at 18-25° C. for 4-8 hours.

11. The method according to any one of claims 1 to 10, characterized in that The yield of the obtained glucose was 95% or more, the purity of the obtained glucose was 85% or more, the yield of the obtained mannose was 95% or more, and the purity of the obtained mannose was 85% or more.

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