Preparation method of maltotetraose aqueous solution with concentration of more than 50%

Abstract

The invention discloses a preparation method of a maltotetraose aqueous solution with the concentration larger than 50%, and the preparation method comprises the following steps: S1, conversion: converting maltodextrin to obtain a maltotetraose conversion solution; s2, degerming: carrying out first filtration on the obtained maltotetraose conversion liquid to obtain first filtered clear liquid; s3, decoloring: decoloring the obtained first filtered clear liquid to obtain a decolored liquid; s4, removing maltodextrin, and carrying out second filtration on the obtained decolored solution to obtain a second filtered clear solution; and S5, concentrating: concentrating the obtained second filtered clear liquid under reduced pressure to obtain a maltotetraose aqueous solution with the concentration of more than 50%. According to the method, the maltodextrin is removed from the maltotetraose conversion liquid by adopting the ultrafiltration membrane for the first time, the content and the yield of the product are effectively improved, and the final yield of the maltotetraose aqueous solution prepared by the method is greater than 90% and the concentration is greater than 50%. The method disclosed by the invention is green and environment-friendly, low in operation and maintenance cost and simple in operation process.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and in particular relates to a method for preparing a maltotetraose aqueous solution with a concentration greater than 50%. Background Art

[0002] Maltotetraose (Maltotetraose, chemical formula is C 24 H 42 O 21 ), is a linear maltooligosaccharide, a chain oligosaccharide polymer composed of 4 α-D-pyranose glucose units connected by α-1,4-glycosidic bonds, often referred to as maltotetraose or "G4". Maltotetraose is a new type of functional oligosaccharide with good processing adaptability and unique physiological effects. First of all, it has excellent food processing performance: when maltooligosaccharides are added to baked foods or puffed foods, they can be used as moisture regulators due to their good water retention properties; when added to beverages, they can be used as thickeners due to their high viscosity. Secondly, maltooligosaccharides also have good physiological effects: after the human body ingests maltooligosaccharides, they can be quickly used to synthesize glycogen after enzymatic hydrolysis in the body, so they are not easy to cause blood sugar to rise; after entering the blood, they will not cause an increase in osmotic pressure, so they can be used as a sports drink additive to improve athletes' endurance; yeast and other bacteria are difficult to use, which can slow down their growth rate to a certain extent, thereby effectively preventing tooth decay.

[0003] At present, the preparation methods of maltotetraose include acid preparation method, enzyme preparation method and microbial enzyme catalysis method. Among them, the production process of microbial enzyme catalysis method is more environmentally friendly and less damaging to production equipment, so it is widely used. However, the maltotetraose produced by the maltotetraose generating enzyme catalyzed substrate in the industry is usually a mixture of various oligosaccharides and incompletely converted maltodextrin, among which maltodextrin accounts for a high proportion. The high proportion of maltodextrin in the product will cause subsequent separation difficulties. The traditional separation and purification process of maltotetraose is complicated and the product yield is low; ion exchange resin is required, but a large amount of sewage will be generated during the activation of the resin, which will cause great environmental pressure and high production cost. Therefore, it is very necessary to explore and establish a simple and easy method for preparing maltotetraose with low production cost. Summary of the invention

[0004] The primary purpose of the present invention is to provide a method for preparing a maltotetraose aqueous solution with a concentration greater than 50%. By adopting the method of the present invention, a high-concentration and high-purity maltotetraose aqueous solution can be obtained.

[0005] To this end, the present invention provides the following technical solutions.

[0006] One aspect of the present invention provides a method for preparing a maltotetraose aqueous solution having a concentration greater than 50%, the preparation method comprising the following steps:

[0007] S1: conversion, converting maltodextrin to obtain maltotetraose conversion liquid;

[0008] S2: sterilizing, filtering the obtained maltotetraose conversion liquid for the first time to obtain a first filtered clear liquid;

[0009] S3: decolorizing, decolorizing the first filtered clear liquid to obtain a decolorized liquid;

[0010] S4: removing maltodextrin, filtering the obtained decolorized liquid for the second time to obtain a second filtered clear liquid;

[0011] S5: Concentration: The obtained second filtered clear liquid is concentrated under reduced pressure to obtain a maltotetraose aqueous solution with a concentration greater than 50%.

[0012] As a preferred embodiment of the present invention, in step S1, the conversion adopts a microbial catalytic conversion method.

[0013] As a preferred embodiment of the present invention, the conversion conditions are: maltodextrin mass concentration is 20-30%, Escherichia coli concentration is 1-10 g / L, conversion temperature is 55°C, rotation speed is 200-600 rpm, and conversion time is 4-7 hours.

[0014] As a preferred embodiment of the present invention, in step S1, the mass concentration of maltotetraose in the maltotetraose conversion liquid is 9-17%, the mass concentration of maltodextrin is 5-10%, the mass concentration of maltose is 0.5-1.5%, and the mass concentration of glucose is 0.1-1%.

[0015] As a preferred embodiment of the present invention, in step S2, the first filtration adopts ceramic microfiltration membrane filtration, the pore size of the ceramic microfiltration membrane is 0.01-0.2 um, and the operating pressure is 0.1-0.5 MPa.

[0016] As a preferred embodiment of the present invention, in step S3, the decolorization is carried out by using activated carbon, the amount of activated carbon added is 0.5-1% of the volume of the first filtered clear liquid, and the decolorization temperature is 30-90°C.

[0017] As a preferred embodiment of the present invention, in step S4, the second filtration is carried out by ultrafiltration membrane filtration, the molecular weight of the ultrafiltration membrane is 5 to 20 KDa, and the operating pressure is 1.5 to 4.0 MPa.

[0018] As a preferred embodiment of the present invention, in step S5, the reduced pressure concentration conditions are: temperature of 30 to 90° C., operating pressure of -0.01 to -0.1 MPa.

[0019] As a preferred embodiment of the present invention, in step S5, the solid content of the maltotetraose aqueous solution is 70-75%.

[0020] Another aspect of the present invention also provides a maltotetraose aqueous solution with a concentration greater than 50% prepared according to the preparation method as described above.

[0021] By means of the above technical scheme, the present invention has at least the following advantages: the present invention can improve the concentration and yield of the obtained maltotetraose aqueous solution by controlling the pH of the reaction system during the conversion process. In addition, the present invention also uses an ultrafiltration membrane for the first time to remove maltodextrin from the maltotetraose conversion liquid, eliminating the ion exchange resin step, and has the advantages of simple process flow and low cost. In addition, the amount of sewage discharged during the production process is small, which reduces the production cost. The maltotetraose aqueous solution prepared by the method of the present invention has a final yield of more than 90% and a concentration of more than 50%.

[0022] The above description is only an overview of the technical solution of the present invention. In order to more clearly understand the technical means of the present invention and implement it according to the contents of the specification, the preferred embodiments of the present invention are described in detail as follows. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 This is a process flow chart for preparing the maltotetraose aqueous solution of the present invention;

[0024] Figure 2 This is a picture of a finished product obtained according to the preparation method of Example 1 of the present invention;

[0025] Figure 3 The liquid phase detection spectrum of the finished product obtained according to the preparation method of Example 1 of the present invention;

[0026] Figure 4 The liquid phase detection spectrum of the finished product obtained according to the preparation method of Example 2 of the present invention;

[0027] Figure 5 The liquid phase detection spectrum of the finished product obtained according to the preparation method of Example 3 of the present invention;

[0028] Figure 6 The liquid phase detection spectrum of the finished product obtained according to the preparation method of Comparative Example 1 of the present invention;

[0029] Figure 7 This is a liquid phase detection spectrum of the finished product obtained according to the preparation method of Comparative Example 2 of the present invention. DETAILED DESCRIPTION

[0030] In order to make the technical means, creative features, objectives and effects of the present invention easy to understand, the technical solutions in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0031] Unless otherwise specified, the percentage content involved in the present invention refers to mass percentage for solid-liquid mixing and solid-solid mixing, and refers to volume percentage for liquid-liquid mixing.

[0032] Unless otherwise specified, the percentage concentrations referred to in the present invention all refer to the final concentration, which refers to the percentage of the added component in the system after the addition of the component.

[0033] The temperature parameters in the present invention, if not specifically limited, allow for both constant temperature treatment and treatment within a certain temperature range. The constant temperature treatment allows the temperature to fluctuate within the precision range controlled by the instrument.

[0034] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods.

[0035] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0036] Unless otherwise specified, the transmittance detection method in the following embodiments is to detect the transmittance at a wavelength of 430 nm using spectrophotometry.

[0037] Unless otherwise specified, the solid content detection method in the following embodiments is to use a saccharimeter to detect the refractive index of the solution.

[0038] Unless otherwise specified, the microorganisms used in the following examples are Escherichia coli, which are deposited in the microbiological laboratory of the applicant.

[0039] Unless otherwise specified, the HPLC analysis conditions in the embodiments of the present invention are shown in Table 1:

[0040] Table 1 Liquid chromatography and conditions

[0041]

[0042] Embodiment 1:

[0043] A method for separating and purifying maltotetraose comprises the following steps:

[0044] S1: Conversion: Weigh 600g of maltodextrin, add water to make up to 2L, and prepare a 30% maltodextrin solution; add 5g of Escherichia coli to the obtained maltodextrin solution, and transform at 55°C and 300rpm for 4h. During the reaction, 10% sodium hydroxide is used to control the system pH to 6.0 to obtain maltotetraose conversion liquid.

[0045] S2: Sterilization: The maltotetraose conversion liquid obtained in step S1 was passed through a ceramic microfiltration membrane with a pore size of 50 nm at an operating pressure of 0.5 MPa to remove bacteria and macromolecular proteins, and 1.95 L of filtered clear liquid was collected.

[0046] S3: Decolorization: Add activated carbon (the amount added is 0.5% of the volume of the filtered clear liquid) to the filtered clear liquid obtained in step S3 and decolorize at 70°C for 60 minutes. After the decolorization is completed, filter to remove the activated carbon and collect the decolorized liquid. The transmittance of the obtained decolorized liquid is 99% as determined by spectrophotometry.

[0047] S4: Removal of maltodextrin: The decolorized liquid obtained in step S3 is passed through an ultrafiltration membrane with a molecular weight of 10 KDa at an operating pressure of 1.5 MPa to remove maltodextrin in the decolorized liquid, and 1.85 L of ultrafiltration membrane clear liquid is collected.

[0048] S5: Concentration: The ultrafiltration membrane clear liquid obtained in the above step S4 was concentrated under reduced pressure at an operating pressure of -0.095 MPa and a temperature of 60°C until the solid content reached 72%, and the concentration was stopped to obtain 522.61 g of maltotetraose aqueous solution.

[0049] like Figure 2 As shown, the obtained maltotetraose aqueous solution has the appearance of a pure, transparent, high-viscosity liquid.

[0050] like Figure 3 As shown, the concentration detected by high performance liquid chromatography is 55.81%, and the yield is calculated to be 92.3%.

[0051] Embodiment 2:

[0052] A method for separating and purifying maltotetraose comprises the following steps:

[0053] S1: Conversion: Weigh 900g of maltodextrin, add water to make up to 3L, and prepare a 30% maltodextrin solution; add 12g of Escherichia coli to the obtained maltodextrin solution, and transform at 55°C and 300rpm for 4h. During the reaction, 10% sodium hydroxide is used to control the system pH to 6.0 to obtain maltotetraose conversion liquid.

[0054] S2: Sterilization: The maltotetraose conversion liquid obtained in step S1 was passed through a ceramic microfiltration membrane with a pore size of 50 nm at an operating pressure of 0.5 MPa to remove bacteria and macromolecular proteins, and 2.90 L of filtered clear liquid was collected.

[0055] S3: Decolorization: Add activated carbon (the amount added is 0.5% of the volume of the filtered clear liquid) to the filtered clear liquid obtained in step S3 and decolorize at 70°C for 60 minutes. After the decolorization is completed, filter to remove the activated carbon and collect the decolorized liquid. The transmittance of the obtained decolorized liquid is 99.5% as determined by spectrophotometry.

[0056] S4: Removal of maltodextrin: The decolorized liquid obtained in step S3 is passed through an ultrafiltration membrane with a molecular weight of 5 KDa at an operating pressure of 1.0 MPa to remove maltodextrin in the decolorized liquid, and 2.78 L of ultrafiltration membrane clear liquid is collected.

[0057] S5: Concentration: The ultrafiltration membrane clear liquid obtained in step S4 was concentrated under reduced pressure at an operating pressure of -0.095 MPa and a temperature of 65°C until the solid content reached 73%, and then the concentration was stopped to obtain 837.61 g of maltotetraose aqueous solution, which appeared to be a pure, transparent, high-viscosity liquid.

[0058] like Figure 4 As shown, the concentration detected by high performance liquid chromatography is 52.07%, and the calculated yield is 92.60%.

[0059] Embodiment 3:

[0060] A method for separating and purifying maltotetraose comprises the following steps:

[0061] S1: Conversion: Weigh 15 kg of maltodextrin, add water to make up to 50 L, and prepare a 30% maltodextrin solution; add 150 g of Escherichia coli to the obtained maltodextrin solution, and transform at 55°C and 400 rpm for 4 hours. During the reaction, 10% sodium hydroxide is used to control the system pH to 6.0 to obtain maltotetraose conversion liquid.

[0062] S2: Sterilization: The maltotetraose conversion liquid obtained in step S1 was passed through a ceramic microfiltration membrane with a pore size of 50 nm at an operating pressure of 0.5 MPa to remove bacteria and macromolecular proteins, and 48.7 L of filtered clear liquid was collected.

[0063] S3: Decolorization: Add activated carbon (the amount added is 0.5% of the volume of the filtered clear liquid) to the filtered clear liquid obtained in step S3 and decolorize at 70°C for 60 minutes. After the decolorization is completed, filter to remove the activated carbon and collect the decolorized liquid. The transmittance of the obtained decolorized liquid is 99.8% as determined by spectrophotometry.

[0064] S4: Removal of maltodextrin: The decolorized liquid obtained in step S3 is passed through an ultrafiltration membrane with a molecular weight of 10 KDa at an operating pressure of 1.5 MPa to remove maltodextrin in the decolorized liquid, and 47.6 L of ultrafiltration membrane clear liquid is collected.

[0065] S5: Concentration: The ultrafiltration membrane clear liquid obtained in step S4 was concentrated under reduced pressure at an operating pressure of -0.095 MPa and a temperature of 75°C until the solid content reached 72%, and then the concentration was stopped to obtain 13.56 kg of maltotetraose aqueous solution, which appeared to be a pure, transparent, high-viscosity liquid.

[0066] like Figure 5 As shown, the concentration detected by high performance liquid chromatography is 51.88%, and the yield is calculated to be 93.30%.

[0067] Comparative Example 1:

[0068] The only difference from Example 1 is that the conversion process in step S1 does not control the system pH to 6.0, and the rest is consistent with Example 1. The specific steps are as follows:

[0069] S1: Conversion: Weigh 600g of maltodextrin, add water to make up to 2L, and prepare a 30% maltodextrin solution; add 5g of Escherichia coli to the obtained maltodextrin solution, and convert at 55°C and 300rpm for 4h. The pH of the system is not controlled during the reaction process to obtain a maltotetraose conversion solution.

[0070] S2: Sterilization: The maltotetraose conversion liquid obtained in step S1 was passed through a ceramic microfiltration membrane with a pore size of 50 nm at an operating pressure of 0.5 MPa to remove bacteria and macromolecular proteins, and 1.97 L of filtered clear liquid was collected.

[0071] S3: Decolorization: Add activated carbon (the amount added is 0.5% of the volume of the filtered clear liquid) to the filtered clear liquid obtained in step S3 and decolorize at 70°C for 60 minutes. After the decolorization is completed, filter to remove the activated carbon and collect the decolorized liquid. The transmittance of the obtained decolorized liquid is 98.6% as determined by spectrophotometry.

[0072] S4: Removal of maltodextrin: The decolorized liquid obtained in step S3 is passed through an ultrafiltration membrane with a molecular weight of 10 KDa at an operating pressure of 1.5 MPa to remove maltodextrin in the decolorized liquid, and 1.90 L of ultrafiltration membrane clear liquid is collected.

[0073] S5: Concentration: The ultrafiltration membrane clear liquid obtained in the above step S4 was concentrated under reduced pressure at an operating pressure of -0.095 MPa and a temperature of 60°C until the solid content reached 72%, and the concentration was stopped to obtain 488.61 g of maltotetraose aqueous solution.

[0074] like Figure 6 As shown, the concentration was 39.58% as detected by HPLC analysis, and the yield was 80.46% as calculated.

[0075] Comparative Example 2:

[0076] The only difference from Example 1 is that in step S4, an organic nanofiltration membrane is used to replace the ultrafiltration membrane. The rest is consistent with Example 1. The specific steps are as follows:

[0077] S1: Conversion: Weigh 600g of maltodextrin, add water to make up to 2L, and prepare a 30% maltodextrin solution; add 5g of Escherichia coli to the obtained maltodextrin solution, and transform at 55°C and 300rpm for 4h. During the reaction, 10% sodium hydroxide is used to control the system pH to 6.0 to obtain maltotetraose conversion liquid.

[0078] S2: Sterilization: The maltotetraose conversion liquid obtained in step S1 was passed through a ceramic microfiltration membrane with a pore size of 50 nm at an operating pressure of 0.5 MPa to remove bacteria and macromolecular proteins, and 1.96 L of filtered clear liquid was collected.

[0079] S3: Decolorization: Add activated carbon (the amount added is 0.5% of the volume of the filtered clear liquid) to the filtered clear liquid obtained in step S3 and decolorize at 70°C for 60 minutes. After the decolorization is completed, filter to remove the activated carbon and collect the decolorized liquid. The transmittance of the obtained decolorized liquid is 99.5%.

[0080] S4: Removal of maltodextrin: The decolorized liquid obtained in step S3 was passed through an organic nanofiltration membrane with a molecular weight of 1000Da at an operating pressure of 2.5 MPa to remove maltodextrin in the decolorized liquid, and 1.92L of nanofiltration membrane clear liquid was collected.

[0081] S5: Concentration: The nanofiltration membrane clear liquid obtained in the above step S4 was concentrated under reduced pressure at an operating pressure of -0.095 MPa and a temperature of 60°C until the solid content reached 73%, and the concentration was stopped to obtain 510.61 g of maltotetraose aqueous solution.

[0082] like Figure 7 As shown, the concentration detected by high performance liquid chromatography is 45.98%, and the calculated yield is 83.46%.

[0083] As can be seen from the above, in the conversion reaction process of Comparative Example 1, 10% sodium hydroxide was not used to control the pH of the system to 6.0, and the concentration of the obtained maltotetraose aqueous solution did not reach 50%, and the yield was less than 90%; in Comparative Example 2, organic membrane nanofiltration was used to replace ultrafiltration to remove maltodextrin, and the concentration of the obtained maltotetraose aqueous solution did not reach 50%, and the yield was less than 90%. Through the comparison of the products, it can be seen that the maltotetraose aqueous solution obtained by the methods of Examples 1 to 3 is significantly better than Comparative Examples 1 and 2 in concentration and yield. Therefore, the preparation method of the present invention is more suitable for the preparation of maltotetraose aqueous solutions with a concentration greater than 50%.

[0084] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Although the present invention has been disclosed as a preferred embodiment as above, it is not used to limit the present invention. Any technician familiar with this profession can make some changes or modifications to equivalent embodiments of equivalent changes by using the methods and technical contents disclosed above without departing from the scope of the technical solution of the present invention. However, any simple modification, equivalent change and modification made to the above embodiments according to the technical essence of the present invention without departing from the content of the technical solution of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. A method for preparing a maltotetraose aqueous solution with a concentration greater than 50%, characterized in that: The preparation method comprises the following steps: S1: conversion, converting maltodextrin to obtain maltotetraose conversion liquid; S2: sterilizing, filtering the obtained maltotetraose conversion liquid for the first time to obtain a first filtered clear liquid; S3: decolorizing, decolorizing the first filtered clear liquid to obtain a decolorized liquid; S4: removing maltodextrin, filtering the obtained decolorized liquid for the second time to obtain a second filtered clear liquid; S5: Concentration: The obtained second filtered clear liquid is concentrated under reduced pressure to obtain a maltotetraose aqueous solution with a concentration greater than 50%.

2. The preparation method according to claim 1, characterized in that: In step S1, the conversion is carried out by microbial catalytic conversion.

3. The preparation method according to claim 2, characterized in that: The conversion conditions are as follows: maltodextrin mass concentration is 20-30%, Escherichia coli concentration is 1-10 g / L, conversion temperature is 50-60° C., rotation speed is 200-600 rpm, and conversion time is 4-7 hours.

4. The preparation method according to claim 1, characterized in that: In step S1, the mass concentration of maltotetraose in the maltotetraose conversion liquid is 9-17%, the mass concentration of maltodextrin is 5-10%, the mass concentration of maltose is 0.5-1.5%, and the mass concentration of glucose is 0.1-1%.

5. The preparation method according to claim 1, characterized in that: In step S2, the first filtration is carried out by filtration using a ceramic microfiltration membrane, the pore size of the ceramic microfiltration membrane is 0.01-0.2 um, and the operating pressure is 0.1-0.5 MPa.

6. The preparation method according to claim 1, characterized in that: In step S3, the decolorization is carried out by using activated carbon, the amount of activated carbon added is 0.5-1% of the volume of the first filtered clear liquid, and the decolorization temperature is 30-90°C.

7. The preparation method according to claim 1, characterized in that: In step S4, the second filtration is carried out by ultrafiltration membrane filtration, the molecular weight of the ultrafiltration membrane is 5-20KDa, and the operating pressure is 1.5-4.0Mpa.

8. The preparation method according to claim 1, characterized in that: In step S5, the reduced pressure concentration conditions are: temperature of 30-90° C., operating pressure of -0.01-0.1 MPa.

9. The preparation method according to claim 1, characterized in that: In step S5, the solid content of the maltotetraose aqueous solution is 70-75%.

10. A maltotetraose aqueous solution having a concentration greater than 50% prepared by the preparation method according to any one of claims 1 to 9.

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