Method for preparing galactose and application thereof

Abstract

The invention relates to the technical field of galactose production, and particularly discloses a method for preparing galactose and application thereof. According to the method for preparing galactose, galactose is prepared through an enzymolysis reaction, a system of the enzymolysis reaction comprises neutral lactase, a reaction medium and lactose or a raw material containing lactose, the reaction medium is a composite salt solution, and the composite salt solution comprises dihydrogen phosphate ions, potassium ions and magnesium ions; the molar ratio of the dihydrogen phosphate ions to the potassium ions to the magnesium ions is (3-8): (3-8): 1, and the pH value of the composite salt solution is 6.5-7.0. According to the method, the lactose raw material can be completely subjected to enzymolysis, and the galactose yield is efficiently increased.

Description

Technical Field

[0001] The invention relates to the technical field of galactose production, in particular to a method for preparing galactose and application thereof. Background Art

[0002] D-galactose is a monosaccharide that can promote the intellectual development of infants and is a synthetic precursor of many drugs. It is widely used in scientific research and production. In the field of food, D-galactose is the direct precursor for the preparation of D-tagatose. D-tagatose is a functional sweetener with a sweetness comparable to that of sucrose and low calories. It has functional properties such as lowering blood sugar, anti-obesity, anti-oxidation, and regulating intestinal flora. The preparation technology of D-galactose has a great impact on the development of D-tagatose. The raw materials used to prepare D-galactose include sugar components such as plant gum, agar, and lactose. Among them, lactose is a disaccharide from milk source containing D-galactose. D-galactose can be generated by degradation. This method is also a common method for the industrial preparation of D-galactose. Lactose is a by-product of deep processing of milk and has the advantages of low price, safe source, and high efficiency.

[0003] There are two methods for preparing D-galactose using lactose as raw material: acid hydrolysis and enzymatic hydrolysis. Acid hydrolysis is basically used in industrial production. It uses inorganic acid as a catalyst to hydrolyze lactose into D-galactose under high temperature and high pressure conditions. This method has low production efficiency, high equipment requirements, many by-products produced by hydrolysis, and incomplete hydrolysis of lactose, which makes the production cost of D-galactose high and the market price expensive. Enzymatic hydrolysis is to hydrolyze lactose into one molecule of glucose and one molecule of D-galactose by lactase. The hydrolysis conditions are mild, the operation is simple, the product purity and yield are high, and the lactase used has long been commercialized and widely used. The development of this method is of great significance to the preparation of D-galactose.

[0004] When lactose is completely hydrolyzed, the theoretical yield of galactose is 50%. Since lactase has a bidirectional effect of hydrolysis and transaside, that is, while hydrolyzing to produce galactose, it can also transfer galactosyl to lactose to produce oligogalactose, which reduces the yield of galactose. At present, the actual industrial production yield of D-galactose is generally around 30%. The specific process of lactase hydrolyzing lactose is as follows: lactase combines with lactose to hydrolyze and first release glucose to form a lactase-galactosyl complex, and then the enzyme transfers the galactosyl to a nucleophilic receptor containing a hydroxyl group (both water and sugar can be used as receptors). When the receptor is water, galactose is generated, and when the receptor is sugar, new sugars are formed, such as trisaccharides, tetrasaccharides, pentasaccharides and other oligogalactoses. The lactose concentration in the hydrolyzate has a great influence on the hydrolysis product. When the lactose concentration is low (<10%), water is more likely to act as an acceptor, and the product is mainly galactose. The higher the lactose concentration, the higher the chance that lactose will act as an acceptor and combine with galactose to form oligosaccharides. In the process of preparing galactose using lactose as raw material, although the galactose conversion rate is higher at low lactose concentration, the galactose yield is lower due to the low lactose concentration, resulting in higher production costs; high lactose concentration is conducive to increasing the galactose yield, but under high lactose concentration conditions, lactose hydrolysis is more likely to undergo transamination to generate oligosaccharides, which will also reduce the galactose yield. Therefore, increasing the galactose yield at high lactose concentration is a key technical difficulty in preparing galactose.

[0005] However, currently, most of the research in the field of lactose hydrolysis is aimed at improving the yield of galacto-oligosaccharides. Generally, the yield of galacto-oligosaccharides is improved by combining process optimization under conditions of higher lactose concentrations. However, there are relatively few studies on the preparation technology and yield improvement of D-galactose, and it has not yet achieved the feasibility of industrial application in terms of technology and economy. Therefore, it is necessary to further study the production method of D-galactose. Summary of the invention

[0006] One of the objects of the present invention is to provide a method for preparing galactose in which raw materials can be fully utilized and galactose production efficiency is high.

[0007] The invention provides a method for preparing galactose. The galactose is prepared by enzymatic hydrolysis reaction. The system of the enzymatic hydrolysis reaction comprises neutral lactase, a reaction medium and lactose or a raw material containing lactose. The reaction medium is a composite salt solution. The composite salt solution comprises dihydrogen phosphate ions, potassium ions and magnesium ions. The molar ratio of the dihydrogen phosphate ions, potassium ions and magnesium ions is (3-8): (3-8): 1. The pH value of the composite salt solution is 6.5-7.0.

[0008] The invention reduces the transaglycosation of lactase and the generation of oligogalactose by specifically adjusting the hydrolysis medium of lactase, thereby obtaining a higher galactose yield, and lactose can be completely hydrolyzed, with high raw material utilization, thus providing a technical reference for the production of D-galactose and the development of the functional sweetener D-tagatose synthesized using D-galactose as a raw material.

[0009] Specifically, the composite salt solution of the present invention is conducive to the occurrence of lactase hydrolysis reaction, can increase the hydrolysis rate of lactose, and shorten the hydrolysis time. The present invention has found that magnesium ions are conducive to improving the overall activity of lactase, dihydrogen phosphate can help reduce the production of oligomeric galactose, and potassium ions can increase the hydrolysis rate of lactase. By combining with dihydrogen phosphate, the conversion rate of galactose can be improved as a whole, and the hydrolysis time can be shortened. When these three specific ions are matched in a specific ratio, enzymolysis with neutral lactase under pH 6.5-7.0 can achieve the effect of sufficient enzymolysis of raw materials and efficient acquisition of galactose as a whole.

[0010] The present invention preferably uses neutral lactase with high enzymatic activity in a neutral environment for enzymolysis. Neutral lactase is commercially available, and its preparation sources include but are not limited to Kluyveromyces lactis, Kluyveromyces fragilis, Bifidobacterium and the like.

[0011] In the present invention, the raw materials containing lactose include but are not limited to whey permeate (powder or liquid), lactose concentrate, cow's milk and any other raw materials containing lactose.

[0012] In the method of the present invention, preferably, the molar ratio of the dihydrogen phosphate ions, potassium ions and magnesium ions is 5:5:1.

[0013] In the method of the present invention, in the enzymatic hydrolysis reaction system, the concentration of lactose in the reaction medium is 15-60% (W / V); preferably 30-50% (W / V).

[0014] Lactose concentration has a great influence on the transaglycosylation of lactase. When the lactose concentration is low, the yield of galactose produced by hydrolysis is high and the content of oligogalactose is low. When the lactose concentration is higher than 10%, as the lactose concentration increases, the more favorable the transaglycosylation reaction is, the higher the conversion rate of galactose and lactose polymerization to generate oligogalactose. When the lactose concentration reaches more than 50%, the solubility of lactose is close to saturation, lactose crystals will appear, and heating is required for full dissolution. At the same time, the viscosity of the feed liquid will increase significantly, making the industrial production operation difficult and the production cost increased. Therefore, the lactose concentration is preferably below 50%.

[0015] The reaction medium in the enzymatic reaction system of the present invention can efficiently produce galactose under the condition of high lactose concentration, avoiding the problem of low galactose yield and high production cost caused by increasing the galactose conversion rate by reducing the lactose concentration.

[0016] In the method of the present invention, based on the mass of lactose, the added amount of lactase is 100-300 U / g; preferably 140-300 U / g.

[0017] In the method of the present invention, the concentration of the composite salt solution is 0.1-0.2 mol / L, preferably 0.2 mol / L.

[0018] The concentration of the total substance (excluding lactose raw materials and enzymes) in the composite salt solution of the present invention is appropriate, which can ensure that the regulating effect on lactase is fully exerted, so as to achieve the effect of improving the galactose yield.

[0019] In the method of the present invention, the dihydrogen phosphate ions come from one or more of potassium dihydrogen phosphate and sodium dihydrogen phosphate; the potassium ions come from one or more of potassium chloride, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium sulfate, and potassium hydroxide; the magnesium ions come from one or more of magnesium chloride and magnesium sulfate; preferably, the dihydrogen phosphate ions come from potassium dihydrogen phosphate.

[0020] In the method of the present invention, the temperature of the enzymatic hydrolysis reaction is 35-40°C, and the time is 3-9h, preferably 3-6h.

[0021] The method of the invention can completely enzymolyze lactose within a relatively short enzymolysis time to obtain a high galactose yield.

[0022] In the method of the present invention, after the enzymatic hydrolysis reaction is completed, the step of inactivating the enzyme is further included, and the conditions for inactivating the enzyme are: heat treatment at 85-100°C for 5-15 minutes, and more preferably, heat treatment at 95°C for 10 minutes.

[0023] In the method of the present invention, after the enzyme is inactivated, the method further comprises the step of centrifuging and obtaining the supernatant to separate and purify galactose.

[0024] After the enzyme is inactivated in the present invention, a mixed sugar solution containing galactose, glucose, etc. can be obtained. In order to further obtain a purer galactose product, the galactose in the mixed sugar solution can be further separated. The separation of galactose can be carried out by conventional methods in the art. For example, desalting and decolorization treatment can be carried out first, including but not limited to activated carbon decolorization, ion exchange, nanofiltration, electrodialysis and other treatments, to remove the color and salt components in the hydrolyzate (supernatant). Then, concentration treatment is carried out, including but not limited to evaporation concentration, nanofiltration membrane concentration, etc. Finally, galactose is obtained by chromatographic separation. Preferably, chromatographic separation is carried out by simulated moving bed separation, and the chromatographic separation resin used includes but is not limited to Ca 2+ Type, K + Type, etc.

[0025] As a specific embodiment, the specific preparation process of galactose of the present invention is as follows: 1. Prepare the reaction medium composite salt solution. Weigh dihydrogen phosphate, potassium salt and magnesium salt to prepare a 0.1-0.2 mol / L salt solution with a pH of 6.5-7.0. The molar ratio of dihydrogen phosphate ion, potassium ion and magnesium ion in the salt solution is (3-8): (3-8): 1. Stir to fully dissolve.

[0026] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose or any raw material containing lactose to prepare a solution with a lactose concentration of 15-60%, and stir or heat and stir to fully dissolve the lactose.

[0027] 3. Enzymatic hydrolysis: add lactase at 100-300U / g lactose and perform enzymatic hydrolysis at 35-40℃ for 3-9h.

[0028] 4. Inactivate the enzyme, heat treat at 85-100°C for 5-15 min, centrifuge at 5000 rpm for 15 min to remove the precipitate, and obtain the supernatant containing galactose.

[0029] The supernatant can be further separated and purified: first, the supernatant obtained by centrifugation is desalted and decolorized, then concentrated to 50-60% Brix, and then chromatographically separated to obtain galactose solution, glucose separation solution and other component sugar separation solutions.

[0030] The present invention also provides application of the method in preparing tagatose.

[0031] The galactose solution prepared by enzymatic hydrolysis of the present invention can be further used to prepare crystalline galactose products through concentration, crystallization and drying, and can also be used as an intermediate raw material for producing tagatose, galacto-oligosaccharides, etc. The byproduct glucose separation solution and other mixed component separation solutions of the present invention can be used for other purposes.

[0032] The beneficial effects of the present invention are at least: The present invention provides a method for preparing galactose, which can completely hydrolyze lactose (lactose conversion rate 100%) in a shorter enzymatic hydrolysis time, increase the galactose yield (greater than 40%), and reduce the galacto-oligosaccharide yield to less than 10%. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 The figure is a schematic diagram of the preparation process of galactose solution of the present invention.

[0034] Figure 2 This is the liquid phase detection spectrum of the supernatant of Example 1 of the present invention.

[0035] Figure 3 The results are as follows: 30% lactose concentration, 140U / g enzyme dosage, hydrolysis time of galactose yield and content. DETAILED DESCRIPTION

[0036] The preferred embodiments of the present invention will be described in detail below in conjunction with examples. It should be understood that the following examples are provided only for the purpose of illustration and are not intended to limit the scope of the present invention. Those skilled in the art may make various modifications and substitutions to the present invention without departing from the purpose and spirit of the present invention.

[0037] The experimental methods used in the following examples are conventional methods unless otherwise specified. The materials, reagents, etc. used in the following examples are commercially available or prepared according to conventional methods in the art unless otherwise specified.

[0038] In the specific embodiments of the present invention, neutral lactase is purchased from Kerry Company. Those skilled in the art may also use other known neutral lactases that can enzymolyze lactose.

[0039] In the specific implementation of the present invention, the components in the supernatant are separated by a chromatographic column and detected by a differential refractive index detector. The liquid chromatography detection conditions of the supernatant are: chromatographic column Sugar-PakTM I WAT085188 (6.5mm×300mm), column temperature 90°C, differential refractive index detector temperature 35°C, pure water mobile phase, flow rate 0.4ml / min, injection concentration 0.5%, injection volume 20 microliters.

[0040] Example 1

[0041] This embodiment provides a process for preparing galactose solution, and the schematic diagram of the process is shown in Figure 1 , as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0042] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95℃ for 10min, centrifuge at 5000rpm for 15min to remove the precipitate, and obtain the supernatant. The obtained supernatant is tested by liquid chromatography, and the spectrum is shown in Figure 2 .

[0043] The supernatant can be further separated and purified: the supernatant obtained by centrifugation is desalted (conductivity <20μs / cm) and decolorized (until clear and transparent) through ion exchange (filler resin model anion resin LX-360, cation resin LX-150), then evaporated and concentrated to 55% Brix, and then separated by chromatography to obtain a purified galactose solution.

[0044] Example 2

[0045] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0046] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 4 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0047] The supernatant can be further separated and purified using the same method as in Example 1.

[0048] Example 3

[0049] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0050] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 9 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0051] The supernatant can be further separated and purified using the same method as in Example 1.

[0052] Example 4

[0053] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0054] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 300 U / g lactose and perform enzymatic hydrolysis at 37°C for 3 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0055] The supernatant can be further separated and purified using the same method as in Example 1.

[0056] Example 5

[0057] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0058] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 50% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 300 U / g lactose and perform enzymatic hydrolysis at 37°C for 3 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0059] The supernatant can be further separated and purified using the same method as in Example 1.

[0060] Example 6

[0061] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0062] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 50% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 300 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0063] The supernatant can be further separated and purified using the same method as in Example 1.

[0064] Example 7

[0065] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.1 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 3:3:1, and stir to fully dissolve.

[0066] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0067] The supernatant can be further separated and purified using the same method as in Example 1.

[0068] Example 8

[0069] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 8:8:1, and stir to fully dissolve.

[0070] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0071] The supernatant can be further separated and purified using the same method as in Example 1.

[0072] Example 9

[0073] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0074] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 15% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0075] The supernatant can be further separated and purified using the same method as in Example 1.

[0076] Example 10

[0077] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0078] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 60% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0079] The supernatant can be further separated and purified using the same method as in Example 1.

[0080] Embodiment 11

[0081] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium sulfate, and potassium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0082] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0083] The supernatant can be further separated and purified using the same method as in Example 1.

[0084] Example 12

[0085] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0086] 2. Dissolve the whey permeate powder. Use the salt solution prepared in step 1 to dissolve the whey permeate powder, so that the lactose concentration in the solution is 30% (W / V), and stir until it is fully dissolved; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, and centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0087] The supernatant can be further separated and purified using the same method as in Example 1.

[0088] Example 13

[0089] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, dipotassium hydrogen phosphate, and sodium dihydrogen phosphate to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0090] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0091] The supernatant can be further separated and purified using the same method as in Example 1.

[0092] Embodiment 14

[0093] This embodiment provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate, magnesium chloride, potassium sulfate, sodium dihydrogen phosphate, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 5:5:1, and stir to fully dissolve.

[0094] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0095] The supernatant can be further separated and purified using the same method as in Example 1.

[0096] Comparative Example 1 (no salt + 30% lactose)

[0097] This comparative example provides a galactose solution preparation process, which is as follows: 1. Dissolve lactose. Prepare a 30% (W / V) lactose aqueous solution with purified water and stir to fully dissolve. 2. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 3. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0098] The supernatant can be further separated and purified using the same method as in Example 1.

[0099] Comparative Example 2 (sodium salt + 30% lactose)

[0100] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate and disodium hydrogen phosphate, prepare 0.2 mol / L pH 7.0 salt solution, and stir to fully dissolve.

[0101] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a 30% (W / V) lactose aqueous solution, and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0102] The supernatant can be further separated and purified using the same method as in Example 1.

[0103] Comparative Example 3 (sodium salt + 50% lactose)

[0104] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate and disodium hydrogen phosphate, prepare 0.2 mol / L pH 7.0 salt solution, and stir to fully dissolve.

[0105] 2. Dissolving lactose: dissolving lactose with the salt solution prepared in step 1 to prepare a 50% (W / V) lactose aqueous solution, and stirring to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0106] The supernatant can be further separated and purified using the same method as in Example 1.

[0107] Comparative Example 4 (sodium salt + 5% lactose)

[0108] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate and disodium hydrogen phosphate, prepare 0.2 mol / L pH 7.0 salt solution, and stir to fully dissolve.

[0109] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a 5% (W / V) lactose aqueous solution, and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0110] The supernatant can be further separated and purified using the same method as in Example 1.

[0111] Comparative Example 5 (sodium salt + 10% lactose)

[0112] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate and disodium hydrogen phosphate, prepare 0.2 mol / L pH 7.0 salt solution, and stir to fully dissolve.

[0113] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a 10% (W / V) lactose aqueous solution, and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0114] The supernatant can be further separated and purified using the same method as in Example 1.

[0115] Comparative Example 6 (sodium salt + 30% lactose, 24h)

[0116] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate and disodium hydrogen phosphate, prepare 0.2 mol / L pH 7.0 salt solution, and stir to fully dissolve.

[0117] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a 30% (W / V) lactose aqueous solution, and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 24 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0118] The supernatant can be further separated and purified using the same method as in Example 1.

[0119] Comparative Example 7 (sodium salt + magnesium ion)

[0120] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate, disodium hydrogen phosphate and magnesium chloride to prepare a 0.2 mol / L salt solution with a pH of 7.0, wherein the molar ratio of dihydrogen phosphate to magnesium ion in the salt solution is 5:1; stir to fully dissolve.

[0121] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0122] The supernatant can be further separated and purified using the same method as in Example 1.

[0123] Comparative Example 8 (sodium salt + potassium ion)

[0124] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate, disodium hydrogen phosphate and potassium chloride to prepare a 0.2 mol / L salt solution with a pH of 7.0, wherein the molar ratio of dihydrogen phosphate to potassium ion in the salt solution is 5:1; stir to fully dissolve.

[0125] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0126] The supernatant can be further separated and purified using the same method as in Example 1.

[0127] Comparative Example 9 (sodium salt + manganese ion)

[0128] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: weigh sodium dihydrogen phosphate, disodium hydrogen phosphate and manganese chloride to prepare a 0.2 mol / L salt solution with a pH of 7.0, wherein the molar ratio of dihydrogen phosphate to manganese ion in the salt solution is 5:1; stir to fully dissolve.

[0129] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve lactose to prepare a solution with a lactose concentration of 30% (W / V), and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0130] The supernatant can be further separated and purified using the same method as in Example 1.

[0131] Comparative Example 10 (Dihydrogen Phosphate and Potassium Ion)

[0132] This comparative example provides a galactose solution preparation process, which is as follows: 1. Preparation of reaction medium: Weigh potassium dihydrogen phosphate and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0. The molar ratio of dihydrogen phosphate to potassium ion in the salt solution is 1:1. Stir to fully dissolve.

[0133] 2. Dissolve lactose: Use the salt solution prepared in step 1 to dissolve a lactose solution with a concentration of 30% (W / V), add and stir to fully dissolve; 3. Enzymatic hydrolysis: add neutral lactase at 140 U / g lactose and perform enzymatic hydrolysis at 37°C for 6 h; 4. Inactivate the enzyme, heat treat at 95°C for 10 min, centrifuge at 5000 rpm for 15 min to remove the precipitate and obtain the supernatant.

[0134] The supernatant can be further separated and purified using the same method as in Example 1.

[0135] Comparative Example 11

[0136] This comparative example provides a galactose solution preparation process, which is substantially the same as the method in Example 1, except that when the reaction medium is configured, the molar ratio of dihydrogen phosphate: potassium ion: magnesium ion in the salt solution is 4.5:4.5:2.

[0137] Comparative Example 12

[0138] This comparative example provides a galactose solution preparation process, which is basically the same as the method in Example 1, except that, in step 1, the reaction medium is prepared by weighing potassium dihydrogen phosphate, calcium chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: calcium ion in the salt solution is 5:5:1, and stirring is performed to fully dissolve.

[0139] Comparative Example 13

[0140] This comparative example provides a galactose solution preparation process, which is basically the same as the method in Example 1, except that, in step 1, the reaction medium is prepared by weighing potassium dihydrogen phosphate, manganese chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: manganese ion in the salt solution is 5:5:1, and stirring is performed to fully dissolve.

[0141] Comparative Example 14

[0142] This comparative example provides a galactose solution preparation process, which is basically the same as the method in Example 1, except that, in step 1, the reaction medium is prepared by weighing potassium dihydrogen phosphate, zinc chloride, and sodium hydroxide (for pH adjustment) to prepare a 0.2 mol / L salt solution with a pH of 7.0, so that the molar ratio of dihydrogen phosphate: potassium ion: zinc ion in the salt solution is 5:5:1, and stirring is performed to fully dissolve.

[0143] Comparative Example 15

[0144] This comparative example provides a galactose solution preparation process, which is substantially the same as the method in Example 1, except that the pH value of the reaction medium is adjusted to 4.5; and acid lactase is used for enzymolysis at a temperature of 55°C.

[0145] Comparative Example 16

[0146] This comparative example provides a process for preparing galactose solution, and the preparation method thereof can be found in Example 1 of CN 102876752 A.

[0147] Experimental example

[0148] In this experimental example, liquid phase detection was performed on the supernatant obtained after enzyme inactivation and centrifugation in each embodiment and comparative example, and various physical and chemical indicators were compared and analyzed.

[0149] The comparative analysis indicators mainly include lactose concentration, lactose hydrolysis time, lactose content, galactose content, galacto-oligosaccharide content in the supernatant, galactose yield, galacto-oligosaccharide yield. The calculation method of galactose / galacto-oligosaccharide yield is galactose or galacto-oligosaccharide content / lactose content (W / W) × 100%. The results are shown in Table 1.

[0150] Table 1

[0151] It can be obtained from the comparative analysis of the comparative examples and the embodiments that when the lactose concentration is low (<10%), the galactose yield is high, but due to the low galactose content in the hydrolyzate, only below 5%, especially when the lactose concentration is 5%, the galactose content in the hydrolyzate is only about 2%, which will greatly increase the concentration cost. When the lactose concentration increases (≥30%), it can be seen from Comparative Examples 2 and 3 that as the lactose concentration increases, the yield of oligogalactose increases. Although the galactose content in the hydrolyzate increases, the overall yield decreases. When the hydrolysis time is extended to 24h (Comparative Example 6), the galactose yield is significantly improved, which may be due to the further hydrolysis of oligogalactose. For the hydrolysis of lactose by acid lactase, Comparative Examples 15 and 16 show that oligogalactose is also produced, and at the same hydrolysis time, the galactose yield is not as high as that of neutral lactase. Different salts have different effects on lactase hydrolysis. When dihydrogen phosphate and sodium salt act on lactase, the galactose content produced is lower than that of dihydrogen phosphate and potassium salt. Potassium salt is conducive to the hydrolysis of lactose to produce galactose. Magnesium ions are conducive to improving the activity of lactase and promoting the occurrence of hydrolysis and transamination reactions. When dihydrogen phosphate, potassium ions and magnesium ions act on lactase in combination, especially when the molar ratio is 5:5:1, 30% lactose concentration is added to 140U / g lactose. After only 6h of hydrolysis, the galactose yield reaches 46.6%; if the enzyme amount is further increased, when 300U / g lactose is added to lactase, the galactose yield reaches 43.3% after 3h of hydrolysis, which can greatly shorten the enzymatic hydrolysis time. When the lactose concentration is further increased to 50%, when 300U / g lactose is added to lactase, the galactose yield is above 40% after 3-6h of hydrolysis. It can be seen that the specific enzymatic hydrolysis medium of the present invention is conducive to the hydrolysis of lactose to produce galactose, can improve the lactose hydrolysis rate, and increase the galactose yield to more than 40%, and the hydrolysis time is significantly shortened compared with the enzymatic hydrolysis medium solution with water.

[0152] The present invention also compares the galactose yield and content of 30% lactose concentration (different dissolving media: water or the reaction medium of Example 1 of the present invention) hydrolyzed at 140 U / g enzyme for different times. The specific results are shown in Figure 3 It can be seen that the composite salt medium of the present invention is conducive to promoting the hydrolysis reaction of lactase to obtain more galactose. The yield of galactose obtained by hydrolysis under the composite salt medium of the present invention is significantly higher than that of water medium. After 6 hours of hydrolysis, the galactose content in the hydrolyzate tends to be balanced between 10-13%. The high galactose content in the hydrolyzate is conducive to subsequent separation and purification.

[0153] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements may be made thereto based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

Claims

1. A method for preparing galactose, wherein galactose is prepared by enzymatic hydrolysis, wherein the enzymatic hydrolysis system comprises neutral lactase, a reaction medium and lactose or a raw material containing lactose, wherein: The reaction medium is a composite salt solution, which includes dihydrogen phosphate ions, potassium ions and magnesium ions; the molar ratio of the dihydrogen phosphate ions, potassium ions and magnesium ions is (3-8): (3-8): 1, and the pH value of the composite salt solution is 6.5-7.

0.

2. The method for preparing galactose according to claim 1, characterized in that The molar ratio of the dihydrogen phosphate ions, potassium ions and magnesium ions is 5:5:

1.

3. The method for preparing galactose according to claim 1 or 2, characterized in that: In the enzymatic hydrolysis reaction system, the concentration of lactose in the reaction medium is 15-60% (W / V); preferably 30-50% (W / V); And / or, based on the mass of lactose, the added amount of lactase is 100-300 U / g; preferably 140-300 U / g.

4. The method for preparing galactose according to any one of claims 1 to 3, characterized in that: The concentration of the composite salt solution is 0.1-0.2 mol / L, preferably 0.2 mol / L.

5. The method for preparing galactose according to any one of claims 1 to 4, characterized in that: The dihydrogen phosphate ions come from one or more of potassium dihydrogen phosphate and sodium dihydrogen phosphate; the potassium ions come from one or more of potassium chloride, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium sulfate, and potassium hydroxide; the magnesium ions come from one or more of magnesium chloride and magnesium sulfate; preferably, the dihydrogen phosphate ions come from potassium dihydrogen phosphate.

6. The method for preparing galactose according to any one of claims 1 to 5, characterized in that: The temperature of the enzymatic hydrolysis reaction is 35-40°C, and the time is 3-9 hours, preferably 3-6 hours.

7. The method for preparing galactose according to any one of claims 1 to 6, characterized in that: After the enzymatic hydrolysis reaction is completed, the step of inactivating the enzyme is further included, and the conditions for inactivating the enzyme are: heat treatment at 85-100° C. for 5-15 minutes.

8. The method for preparing galactose according to claim 7, characterized in that: The enzyme inactivation condition is: heat treatment at 95°C for 10 min.

9. The method for preparing galactose according to any one of claims 7 to 8, characterized in that: After the enzyme is inactivated, the method further comprises the step of centrifuging and taking the supernatant to separate and purify galactose.

10. Use of the method according to any one of claims 1 to 9 in the preparation of tagatose.

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