Preparation method of glucuronic acid

Abstract

The invention discloses a preparation method of glucuronic acid, which comprises the following steps: preparing a glucuronic acid solution, carrying out first concentration on the glucuronic acid solution until no water is evaporated to obtain glucuronic acid solid powder, adding the glucuronic acid solid powder into a high-concentration short-chain alcohol solution, heating to 40-60 DEG C, re-dissolving, carrying out first filtration to obtain a clear liquid, and drying to obtain the glucuronic acid. And carrying out second concentration on the clear liquid, cooling to 10-20 DEG C at a cooling rate of 1-5 DEG C / h when crystals are separated out, and carrying out second filtration and drying to obtain glucuronic acid dry powder. The preparation method of glucuronic acid disclosed by the invention has the advantages of simplicity, high efficiency, few byproducts and suitability for industrial production.

Description

Technical Field

[0001] The invention relates to the technical field of organic matter preparation, and in particular to a method for preparing glucuronic acid. Background Art

[0002] Glucuronic acid, also known as glucuronic acid, is a compound formed by the oxidation of the primary alcohol hydroxyl group of glucose to a carboxyl group. It is widely distributed in animals and plants. In plants, it exists in gums such as gum arabic in the form of monouronic acid and is an important component of pectin, mucus and higher polysaccharides. In animals, glucuronic acid forms a complex with glucosinolates and is stored in tissues. The molecular formula of glucuronic acid is C 6 H 10 O 7 , relative molecular weight 194.14, powder or white needle crystals. Glucuronic acid aqueous solution is unstable, easy to form 3,6-glucuronic acid lactone and in a state of tautomerization equilibrium. D-glucuronic acid is also easy to decarboxylate and generate CO when heated and in the presence of strong acid. 2 , furan and other cracking products. Glucuronic acid can combine with toxins in the body to form non-toxic glucuronic acid conjugates and be excreted, so it has the effect of protecting the liver and detoxifying. It is often used for hepatitis, cirrhosis, food and drug poisoning. Glucuronic acid is also an important component of connective tissue and can be used to treat arthritis and collagen diseases.

[0003] In the prior art, glucuronic acid is mostly produced by hydrolyzing gum arabic, such as the solution described in document CN112876520A. However, this method has a complicated hydrolysis process, many by-products, and complex separation, and has strict requirements on the separation process, which makes it difficult to achieve industrial production.

[0004] Therefore, the present invention urgently needs to provide a method for preparing glucuronic acid which is simple, efficient, has few by-products and is suitable for industrial production. Summary of the invention

[0005] In view of this, the purpose of the present invention is to provide a method for preparing glucuronic acid, which overcomes the technical problems in the prior art of preparing glucuronic acid by hydrolysis of gum arabic, such as complex hydrolysis process, large number of by-products, complex separation, and difficulty in realizing industrial production.

[0006] In order to achieve the above object, the present invention provides a method for preparing glucuronic acid, comprising the following steps:

[0007] preparing a glucuronic acid solution;

[0008] The glucuronic acid solution is first concentrated until no water is evaporated to obtain a glucuronic acid solid powder;

[0009] Adding the glucuronic acid solid powder into a high-concentration short-chain alcohol solution, heating to 40-60° C. to redissolve, and filtering to obtain a clear solution;

[0010] The clear liquid is concentrated for the second time, and when crystals are precipitated, the temperature is cooled to 10-20° C. at a cooling rate of 1-5° C. / h, and the liquid is filtered and dried for the second time to obtain glucuronic acid dry powder.

[0011] The preparation method of glucuronic acid provided by the present invention first prepares a glucuronic acid solution, the preparation process is simple and efficient, there are no other side reactions, the obtained glucuronic acid solution is then evaporated and dissolved in alcohol to remove impurities, the purity of glucuronic acid is further improved, and finally high-purity glucuronic acid is obtained by cooling and crystallization. The preparation method of glucuronic acid disclosed in the present invention has the characteristics of being simple and efficient, having few by-products and low cost, and is a method suitable for industrial production of glucuronic acid, which lays the foundation for large-scale industrial production. After testing, the content of glucuronic acid in the glucuronic acid product prepared by the method of the present invention can reach more than 99%. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Attached Figure 1 Described is the liquid chromatography spectrum of the glucuronic acid dry powder in Example 1;

[0013] Attached Figure 2 Described is the liquid chromatography spectrum of the glucuronic acid dry powder in Comparative Example 1. DETAILED DESCRIPTION

[0014] In order to make the technical problems, technical solutions and beneficial effects to be solved by the present invention more clearly understood, the present invention is further described in detail below in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0015] The present invention provides a method for preparing glucuronic acid, comprising the following steps:

[0016] preparing a glucuronic acid solution;

[0017] The glucuronic acid solution is first concentrated until no water is evaporated to obtain a glucuronic acid solid powder;

[0018] Adding the glucuronic acid solid powder into a high-concentration short-chain alcohol solution, heating to 40-60° C. to redissolve, and filtering to obtain a clear solution;

[0019] The clear liquid is concentrated for the second time, and when crystals are precipitated, the temperature is cooled to 10-20° C. at a cooling rate of 1-5° C. / h, and the liquid is filtered and dried for the second time to obtain glucuronic acid dry powder.

[0020] The present invention first prepares a glucuronic acid solution. In the present invention, the preparation of the glucuronic acid solution includes: adding a bacterial cell containing inositol oxidase to an inositol feed liquid for conversion to obtain a conversion liquid; filtering the conversion liquid through a third filtration to remove the bacterial cell to obtain a glucuronic acid solution; wherein, preferably, the concentration of the inositol feed liquid is 10-100g / L; preferably, the amount of the bacterial cell containing inositol oxidase added is such that the bacterial cell concentration of the system reaches 20-40OD; the temperature of the conversion is preferably 30°C-40°C, the pH is preferably 8.0-9.0, and the time is preferably 4-6h. The present invention uses inositol feed liquid as a raw material to prepare glucuronic acid, and has the advantages of simple process, high production efficiency, few by-products, and high yield.

[0021] Specifically, the third filtration is preferably plate-and-frame filtration or ceramic membrane filtration; the filter membrane pore size of the plate-and-frame filtration is preferably 10-100nm; in some embodiments of the present invention, any value in the range of 10-100nm can be selected as the filter cloth pore size; specifically, the filter cloth pore size of the plate-and-frame filtration can be selected as 10nm, 20nm, 30nm, 40nm, 50nm, 60nm, 70nm, 80nm, 90nm or 100nm. The present invention has no special requirements for the equipment of the plate-and-frame filtration, and has no special requirements for other parameters of the plate-and-frame filtration such as the number of filter layers, filtration pressure, water flow, etc., as long as filtration under the filter cloth pore size of 10-100nm can be achieved. However, it has been verified by experiments that when the filtration pressure is 0.6MPa and the flow rate is 150L / h·㎡, a better effect can be achieved (the plate-and-frame clear liquid is clear and there are no visible impurities). The separation pore size of the ceramic membrane is preferably 20-100nm; in some embodiments of the present invention, any value in the range of 20-100nm can be selected as the separation pore size of the ceramic membrane; specifically, the separation pore size of the ceramic membrane can be selected as 20nm, 30nm, 40nm, 50nm, 60nm, 70nm, 80nm, 90nm, 100nm. Preferably, when the pore size of the ceramic membrane is 50nm, a better production filtration effect can be achieved, and the clear liquid of the ceramic membrane is clear and free of visible impurities. The present invention has no special requirements for the type of ceramic membrane. Exemplarily, the ceramic membrane can be a tubular ceramic membrane or a flat ceramic membrane. The present invention has no special requirements for the main material of the ceramic membrane. Exemplarily, inorganic ceramic materials such as alumina, zirconium oxide, titanium oxide and silicon oxide can be used. It should be understood that the role of plate and frame filtration or ceramic membrane filtration in the present invention is to filter out insoluble impurities in the solution to achieve the purpose of preliminary purification.

[0022] After obtaining the glucuronic acid solution, the present invention performs a first concentration on the glucuronic acid solution until no water is distilled out to obtain a glucuronic acid solid powder. In the present invention, the temperature of the first concentration is preferably 70-90° C., more preferably 80° C.; the vacuum degree is preferably ≤-0.09 MPa. The purpose of concentrating and evaporating the glucuronic acid solution in the present invention is to remove the water-soluble impurities retained in the solution during the preparation of the glucuronic acid solution in the previous step, and to improve the purity of glucuronic acid.

[0023] After obtaining the glucuronic acid solid powder, the present invention adds the glucuronic acid solid powder to a high concentration short-chain alcohol solution, heats it to 40-60°C to redissolve it, and obtains a clear liquid through a first filtration. In the present invention, the short-chain alcohol solution is preferably an ethanol solution with a concentration of 85-95% (v / v) or a methanol solution with a concentration of 85-95% (v / v); the amount of the ethanol solution or the methanol solution added is preferably 1-3 times the mass of the glucuronic acid solid powder. Further preferably, the heating to 40-60°C to redissolve it also includes: stirring at a rate of 80-120rpm. The purpose of dissolving the glucuronic acid solid powder in the present invention is to dissolve the glucuronic acid through short-chain alcohols, and the impurities therein are separated because they cannot be dissolved in short-chain alcohols. In the present invention, the first filtration preferably uses filter paper, and the filter pore size is preferably 30-80um.

[0024] After obtaining the clear liquid, the present invention performs a second concentration on the clear liquid, and when crystals are precipitated, the temperature is lowered to 10-20°C at a cooling rate of 1-5°C / h, and the glucuronic acid powder is obtained by second filtration and drying. In the present invention, the temperature of the second concentration is preferably 50-70°C; the vacuum degree is preferably ≤-0.09MPa; the second filtration preferably uses a sand core funnel, and the filtration pore size is preferably 4.5-15um; the drying temperature is preferably 40-60°C.

[0025] The present invention also includes glucuronic acid, which is prepared according to the above-mentioned method for preparing glucuronic acid, and the glucuronic acid content in the glucuronic acid dry powder is not less than 99wt%.

[0026] In order to better illustrate the technical solution of the present invention, the present invention also provides the following specific examples. It should be understood that the raw materials used in the following examples are all commercially available raw materials unless otherwise specified.

[0027] Example 1

[0028] S1. 60 L of 50 g / L inositol solution was prepared, whole cells containing inositol oxidase were added to make the bacterial concentration of the system reach 30 OD, and the transformation was carried out at 35 ° C and pH = 8.5 for 5 h to obtain 61 L of transformation solution;

[0029] S2. 61L of the conversion solution was filtered through a plate-and-frame filter with a pore size of 50nm at a filtration pressure of 0.6MPa and a flow rate of 150L / h·㎡ to collect 69L of glucuronic acid solution;

[0030] S3. 69L of glucuronic acid solution was concentrated at 70°C and -0.09MPa to obtain 3.2kg of glucuronic acid solid powder;

[0031] S4. Add 6.4 kg of 90% (v / v) ethanol solution to the solid powder of glucuronic acid, heat to 50 ° C to redissolve, and filter through 50 um filter paper to obtain 9.6 L of clear solution;

[0032] S5. Take 9.6L of clear liquid and concentrate it at 50℃ and -0.09MPa. When crystals are precipitated, cool it to 15℃ at a cooling rate of 3℃ / h, filter it through a 10um sand core funnel, and dry it at 50℃ to obtain 2.76kg of glucuronic acid dry powder.

[0033] The purity of glucuronic acid detected by liquid chromatography was 99.6wt%.

[0034] Example 2

[0035] S1. 60 L of 10 g / L inositol solution was prepared, whole cells containing inositol oxidase were added to make the bacterial concentration of the system reach 20 OD, and the transformation was carried out at 40 ° C and pH = 9 for 4 h to obtain 61 L of transformation solution;

[0036] S2. Filter 61L of the conversion solution through a ceramic membrane with a pore size of 60nm to collect 68L of glucuronic acid solution;

[0037] S3. 68L of glucuronic acid solution was concentrated at 80°C and -0.095MPa to obtain 0.66kg of glucuronic acid solid powder;

[0038] S4. 1.98 kg of 95% (v / v) ethanol solution was added to the solid powder of glucuronic acid, heated to 60 ° C and stirred at a rate of 120 rpm to redissolve, and filtered through 80 um filter paper to obtain 2.8 L of clear solution;

[0039] S5. Take 2.8L of clear liquid and concentrate it at 60℃ and -0.095MPa. When crystals are precipitated, cool it to 20℃ at a cooling rate of 1℃ / h, filter it through a 4.5um sand core funnel, and dry it at 60℃ to obtain 0.59kg of glucuronic acid dry powder.

[0040] The purity of glucuronic acid detected by liquid chromatography was 99.8wt%.

[0041] Example 3

[0042] S1. 60 L of 100 g / L inositol solution was prepared, whole cells containing inositol oxidase were added to make the bacterial concentration of the system reach 40 OD, and the transformation was carried out at 30 ° C and pH = 9 for 6 h to obtain 61 L of transformation solution;

[0043] S2. Filter 61L of the conversion solution through a ceramic membrane with a pore size of 20nm to collect 74L of glucuronic acid solution;

[0044] S3. 74L of glucuronic acid solution was concentrated at 90°C and -0.10MPa to obtain 6.5kg of glucuronic acid solid powder;

[0045] S4. 6.5 kg of 85% (v / v) methanol solution was added to the glucuronic acid solid powder, heated to 40 ° C and stirred at a rate of 80 rpm to redissolve, and filtered through a 30 um filter paper to obtain 11.3 L of clear solution;

[0046] S5. Take 11.3L of clear liquid and concentrate it at 70℃ and -0.10MPa. When crystals are precipitated, cool it to 10℃ at a cooling rate of 5℃ / h, filter it through a 15um sand core funnel, and dry it at 40℃ to obtain 5.8kg of glucuronic acid dry powder.

[0047] The purity of glucuronic acid detected by liquid chromatography was 99.7wt%.

[0048] Comparative Example 1

[0049] The "90% (v / v) ethanol solution" in step S4 of Example 1 was replaced with an equal amount of "80% (v / v) ethanol solution", and other conditions remained unchanged.

[0050] The purity of glucuronic acid detected by liquid chromatography was 92.4 wt %. The reason is that after the concentration of alcohol as a solvent was reduced, the dissolution of glucuronic acid was incomplete, resulting in a decrease in purity.

[0051] Comparative Example 2

[0052] The "cooling at a cooling rate of 3°C / h" in step S5 of Example 2 is adjusted to "cooling at a cooling rate of 8°C / h", and other conditions remain unchanged.

[0053] The purity of glucuronic acid detected by liquid chromatography was 91.2 wt %. The reason was that the cooling speed was too fast, resulting in an incomplete crystallization process and a reduced amount of crystal precipitation.

[0054] Comparative Example 3

[0055] The step S3 in Example 3, "74 L of glucuronic acid solution was concentrated at 90° C. to obtain 6.5 kg of glucuronic acid solid powder" was adjusted to "74 L of glucuronic acid solution was concentrated at 90° C. to obtain 7.0 kg of glucuronic acid solid powder", i.e., part of the water in the glucuronic acid was retained after concentration, and other conditions remained unchanged.

[0056] The purity of glucuronic acid detected by liquid chromatography was 89.5 wt %. The reason was that the product was not evaporated to dryness during the concentration process, so some impurities were not removed, resulting in a lower purity of the final product.

[0057] *Test Case

[0058] Furthermore, the liquid chromatography test conditions in Examples 1-3 of the present invention and Comparative Examples 1-3 are as follows:

[0059] Mobile phase: 10mmol / L formic acid aqueous solution;

[0060] Chromatographic column: calcium column (300*7.7 or similar column);

[0061] Flow rate: 0.5Ml / min;

[0062] Detector: differential detector;

[0063] Column temperature: 55 °C;

[0064] Detector temperature: 45°C;

[0065] Solvent: 10mmol / L formic acid aqueous solution;

[0066] Standard concentration: 1.0 mg / mL (D-glucuronic acid aldehyde [CAS number] 6556-12-3, C 6 H 10 O 7 The content is 99.9%, China Food and Drug Inspection Institute);

[0067] Test sample concentration: 1.0 mg / mL;

[0068] Chromatographic conditions: run with 100% 10 mmol / L formic acid aqueous solution for 30 min.

[0069] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention is described in detail with reference to the aforementioned embodiments, those skilled in the art can still modify the technical solutions described in the aforementioned embodiments or replace some of the technical features therein by equivalents. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for preparing glucuronic acid, characterized in that: The following steps are involved: preparing a glucuronic acid solution; The glucuronic acid solution is first concentrated until no water is evaporated to obtain a glucuronic acid solid powder; Adding the glucuronic acid solid powder into a high-concentration short-chain alcohol solution, heating to 40-60° C. to redissolve, and filtering to obtain a clear solution; The clear liquid is concentrated for the second time, and when crystals are precipitated, the temperature is cooled to 10-20° C. at a cooling rate of 1-5° C. / h, and the liquid is filtered and dried for the second time to obtain glucuronic acid dry powder.

2. The method according to claim 1, characterized in that The method for preparing the glucuronic acid solution comprises: Adding bacteria containing inositol oxidase to the inositol liquid for transformation to obtain a transformation liquid; The transformation liquid is filtered through a third filter to remove the bacterial cells, thereby obtaining a glucuronic acid solution; Wherein, the concentration of the inositol solution is 10-100 g / L; and / or, The amount of the bacteria containing inositol oxidase added is such that the bacterial concentration of the system reaches 20-40 OD.

3. The method according to claim 2, characterized in that The temperature of the conversion is 30°C-40°C, the pH is 8.0-9.0, and the time is 4-6 hours.

4. The method according to claim 2 or 3, characterized in that: The third filtration adopts plate and frame filtration or ceramic membrane filtration; and / or, The pore size of the filter membrane of the plate and frame filtration is 10-100 nm; and / or the separation pore size of the ceramic membrane is 20-100 nm.

5. The method according to claim 1, characterized in that The temperature of the first concentration is 70-90° C., and the vacuum degree is ≤-0.09 MPa.

6. The method according to claim 1, characterized in that The short-chain alcohol solution is an ethanol solution with a concentration of 85-95% (v / v) or a methanol solution with a concentration of 85-95% (v / v); and / or; The added amount of the ethanol solution or methanol solution is 1-3 times the mass of the glucuronic acid solid powder.

7. The method according to claim 1, characterized in that The heating to 40-60°C for re-dissolving also includes: Stirring was carried out at a rate of 80-120 rpm.

8. The method according to claim 1, characterized in that The first filtration uses filter paper with a filtration pore size of 30-80 um; and / or the second filtration uses a sand core funnel with a filtration pore size of 4.5-15 um.

9. The method according to claim 1, characterized in that: The temperature of the second concentration is 50-70°C, and the vacuum degree is ≤-0.09MPa; and / or, The drying temperature is 40-60°C.

10. A glucuronic acid, characterized in that The glucuronic acid powder is prepared by the method for preparing glucuronic acid according to any one of claims 1 to 9, wherein the glucuronic acid content in the glucuronic acid dry powder is not less than 99 wt%.

Images