Preparation method of 3-hydroxybutyric acid betaine salt

Abstract

The invention discloses 3-hydroxybutyric acid betaine salt, which has the following molecular structure: # imgabs0 #, in which Mn < + > is a metal ion selected from Na < + >, K < + >, Mg < 2 + > and Ca < 2 + >; n + is the number of charges and represents 1-2 positive charges. The betaine 3-hydroxybutyrate salt is used as a food additive and a pharmaceutical active ingredient for relieving or preventing fatty liver. The invention also discloses a preparation method of the 3-hydroxybutyric acid betaine salt.

Description

Technical Field

[0001] The invention belongs to the field of nutritional additives, and in particular relates to 3-hydroxybutyric acid betaine salt, a preparation method thereof and use thereof. Background Art

[0002] 3-Hydroxybutyric acid is one of the three ketone bodies produced by fat metabolism in the human body. It has multiple physiological functions and can help the human body to efficiently produce energy in the absence of glucose. 3-Hydroxybutyric acid is produced by fat metabolism in the liver, transports corresponding tissues and organs through blood circulation, and oxidizes in cell mitochondria to generate energy, that is, it can increase the consumption of body fat. K. Clark published the therapeutic effect of 3-hydroxybutyric acid and its salts / ester compounds on patients with liver fat-related diseases in patent document CN 110869012 A. The liver fat in the subjects can be reduced, and it can also be used to treat healthy subjects and subjects with diseases related to increased liver fat. Hu Yanwang disclosed in patent document CN109734575A that 3-hydroxybutyric acid and its salts can be used as nutritional additives or drugs, with great health and medicinal value.

[0003] Studies have found that when running with ketone bodies, the mechanical efficiency of the heart increases by 30%, blood flow increases by 75%, burning fatty acids increases free radicals, and 3-hydroxybutyrate destroys them. During ketosis exercise, even with available muscle glycogen, the burning of intramuscular fat increases 20-fold. This means that the cells ignore the normally preferred glucose reserves and burn more fat.

[0004] Betaine is a nutritional additive that can be used in the pharmaceutical industry. It has the effects of resisting fatty liver and protecting kidneys. It can also be used in health foods and beauty foods. Abdelmalek et al. (2009) conducted a randomized controlled study on 55 patients with non-alcoholic fatty liver disease, of which 34 patients underwent liver tissue biopsy after treatment. It was found that the degree of liver fatty degeneration in the oral betaine group was improved compared with the control group; Saunderson (1990) proved that adding betaine to the diet of growing chickens reduced body fat and increased protein content; Xu Zirong et al. (1998) reported that betaine can increase the content of free carnitine in the liver of chicks, and carnitine helps the β-oxidation of long-chain fatty acids; Xu Zhongnan et al. (2006) fully affirmed the therapeutic effect of betaine on alcoholic fatty liver in rats. Summary of the invention

[0005] As a manufacturer of fine chemicals related to 3-hydroxybutyric acid and betaine, based on their related and identical properties in terms of physiological activity as food additives and active pharmaceutical ingredients, we have conceived a feasible exploration approach for the composite salt formation of the two, and experimentally verified that a composite salt crystal with stable properties was obtained. Specifically, the present invention includes the following technical solutions.

[0006] The first aspect of the present invention provides a 3-hydroxybutyric acid betaine salt having the following molecular structure:

[0007]

[0008] Wherein, n is a natural number 1 or 2; M is an alkali metal or an alkaline earth metal selected from Na, K, Mg and Ca; that is, M n+ is a metal ion selected from Na + , K + Mg 2+ and Ca 2+ ;n + is the charge number, representing 1-2 positive charges,

[0009] The 3-hydroxybutyric acid is selected from D-3-hydroxybutyric acid, L-3-hydroxybutyric acid and DL-3-hydroxybutyric acid, that is, racemic 3-hydroxybutyric acid.

[0010] In one embodiment, the equivalent ratio of betaine to 3-hydroxybutyrate in the 3-hydroxybutyrate betaine salt is 1:0.3-3, for example, 1:0.5-2 or 1:0.8-1.5.

[0011] Preferably, the equivalent ratio of betaine to 3-hydroxybutyrate in the 3-hydroxybutyrate betaine salt is about 1:1.

[0012] It should be understood that when expressing numerical characteristics in this document, the term "approximately" or "around" means that the number represented may have an error range or floating range of ±10%, ±9%, ±8%, ±7%, ±6% or ±5%.

[0013] Preferably, the content of crotonic acid as an impurity in the 3-hydroxybutyric acid betaine salt is not higher than 0.2 wt %, preferably below 0.18 wt %.

[0014] The second aspect of the present invention provides a method for preparing the above-mentioned 3-hydroxybutyric acid betaine salt,

[0015]

[0016] When the salt is a sodium salt, a potassium salt or a magnesium salt, the method comprises the following steps:

[0017] (1) dissolving betaine and 3-hydroxybutyric acid metal salt in a predetermined equivalent ratio, for example, an equivalent ratio (1:1) in water to obtain an aqueous solution;

[0018] (2) heating and stirring the aqueous solution until the system is dissolved and clear, and continuing to stir and react for 1-5 hours;

[0019] (3) Filter while hot to remove insoluble impurities in the system;

[0020] (4) concentrating the filtrate in step (3), for example, by vacuum distillation to substantially remove water and form a composite salt;

[0021] (5) adding an organic solvent to the composite salt obtained in step (4) until crystals are precipitated (crystallization);

[0022] (6) cooling the mixture to a temperature below 10° C. until almost all crystals are precipitated (crystal growth);

[0023] (7) filtering and drying the filter cake to obtain 3-hydroxybutyric acid betaine salt crystals, and

[0024] When the salt is a calcium salt, the method comprises the following steps:

[0025] (1) dissolving betaine and calcium 3-hydroxybutyrate in a predetermined equivalent ratio in water to obtain an aqueous solution;

[0026] (2) heating and stirring the aqueous solution until the system is dissolved and clear, and continuing to stir and react for 1-5 hours;

[0027] (3) Filter while hot to remove insoluble impurities in the system;

[0028] (4) concentrating the filtrate in step (3) until crystals precipitate (crystallization);

[0029] (6) standing and cooling until the crystals are almost completely precipitated (crystal growth);

[0030] (7) filtering and drying to obtain 3-hydroxybutyric acid betaine calcium salt.

[0031] The 3-hydroxybutyric acid metal salt used in the above step (1) is generally a base neutralization reaction product of 3-hydroxybutyric acid.

[0032] The organic solvent described in the above step (5) is generally an organic solvent approved for use by food and drug administration agencies such as the State Food and Drug Administration or FDA, for example, selected from: ethanol, isopropanol, isoamyl alcohol, acetone, ethyl acetate or a mixture of two or more thereof to ensure the safety of the product. Preferably, the organic solvent is acetone or ethyl acetate.

[0033] Preferably, the stirring reaction temperature in the above step (2) is controlled below 65°C, for example, in the range of 55°C-60°C, to avoid the production of crotonic acid as a byproduct from 3-hydroxybutyric acid.

[0034] Preferably, the above method further comprises step (8): recovering the organic solvent by distillation or rectification.

[0035] It should be understood that the term "substantially" herein refers to the vast majority or main part under specific conditions, such as more than 80%, more than 85%, more than 90%, more than 92%, more than 95% or more than 98% of the total.

[0036] The third aspect of the present invention provides the use of the above-mentioned 3-hydroxybutyric acid betaine salt as a food additive and a pharmaceutical active ingredient in food and medicine.

[0037] Specifically, the above-mentioned food is used to reduce or prevent fatty liver or protect kidneys, and the above-mentioned medicine is used to treat fatty liver or protect kidneys.

[0038] The 3-hydroxybutyrate betaine sodium / potassium / magnesium / calcium salt of the present invention has a stable crystal structure and a low impurity content, and can therefore be sold and used as a compound or a fine chemical, for example, as a food additive and a pharmaceutical active ingredient for preparing foods and medicines that have the function of preventing, alleviating or treating fatty liver, or protecting the kidneys. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 The present invention is a process flow chart of an implementation method of preparing 3-hydroxybutyric acid betaine sodium salt.

[0040] Figure 2 It is the HPLC spectrum of 3-hydroxybutyric acid betaine sodium salt prepared in Example 1. DETAILED DESCRIPTION

[0041] The design idea of ​​the present invention is to provide the multifunctional 3-hydroxybutyric acid and betaine to the body together, so as to play a better effect in terms of fat consumption of the human body. The technical scheme adopted is to make 3-hydroxybutyric acid and betaine into a composite salt, and combine the two components into one, which is of great significance for improving human physique and promoting human health as a nutritional additive. At present, there is no product on the market that prepares the above two substances into a composite salt.

[0042] Betaine is an alkaloid with the chemical name N,N,N-trimethylglycine. Its chemical structure is similar to that of amino acids. It is an inner salt and belongs to the quaternary ammonium base class. Therefore, the complex salt needs to have a metal ion that matches the 3-hydroxybutyrate. In order to ensure food safety, the metal ions are alkali metal ions and alkaline earth metal ions Na + , K+ Mg 2+ and Ca 2+ That is, the reaction raw material 3-hydroxybutyric acid metal salt includes sodium salt, potassium salt, magnesium salt and calcium salt. Correspondingly, the 3-hydroxybutyric acid betaine composite salt can be 3-hydroxybutyric acid betaine sodium salt, 3-hydroxybutyric acid betaine potassium salt, 3-hydroxybutyric acid betaine magnesium salt and 3-hydroxybutyric acid betaine calcium salt.

[0043] Herein, the terms "3-hydroxybutyric acid betaine salt" and "3-hydroxybutyric acid betaine complex salt" have the same meaning and can be used interchangeably. For the convenience of description, it can be referred to as "complex salt".

[0044] The composite salt of the present invention at least retains the physiologically active functions of 3-hydroxybutyric acid and betaine, including reducing fatty liver, protecting kidneys, etc. The composite salt can be restored to free 3-hydroxybutyric acid and betaine after being dissolved in water.

[0045] In one embodiment, the equivalent ratio of betaine to 3-hydroxybutyrate in the composite salt can be 1:0.3-3, for example, about 1:1. It is easy for those skilled in the art to understand that since the two components, betaine and 3-hydroxybutyrate, have different effects, the ratio of the two components can be adjusted according to specific uses.

[0046] Experiments have shown that the physical property stability of a composite salt that does not contain a crystallization mother liquor such as crystallization water is higher than that of a composite salt that contains a crystallization mother liquor such as crystallization water. Therefore, in the process of preparing a composite salt, when betaine and 3-hydroxybutyrate undergo a synthesis reaction in an aqueous solution and are combined together by ionic bonds, hydrogen bonds, and van der Waals forces, crystallization is not performed in an aqueous solution to avoid the inclusion of crystallization water, but water is removed and crystallization is performed using a cooled organic solvent such as acetone, and the resulting composite salt does not contain an organic solvent such as acetone.

[0047] It should be understood that in order to ensure the food safety of the composite salt, the organic solvent is an organic solvent approved for use by the State Food and Drug Administration and the FDA, for example, it can be selected from ethanol, isopropanol, isopentanol, acetone, ethyl acetate or a mixture of any proportion of two or more thereof.

[0048] Figure 1 A process flow chart of an embodiment of the present invention for preparing 3-hydroxybutyric acid betaine salt is shown.

[0049] In order to ensure the edible safety of the composite salt, the generation of impurities in the composite salt preparation process is controlled as much as possible. The impurities in the composite salt are mainly crotonic acid (butenoic acid), a byproduct of 3-hydroxybutyric acid, so the composite salt synthesis reaction temperature is controlled below 65°C, thereby effectively controlling the content of crotonic acid in the finished 3-hydroxybutyric acid betaine salt to below 0.2wt%.

[0050] Example

[0051] The present invention is further described below by examples. It should be understood that these examples are only for illustrative purposes, rather than limiting the present invention. Various changes or adjustments made by those skilled in the art based on the present invention should all fall within the scope of protection of the present invention. For example, those skilled in the art use chiral 3-hydroxybutyric acid such as metal salts of D-3-hydroxybutyric acid and betaine to prepare composite salts, and the synthesis process is similar to the process conditions of calcium, magnesium, sodium, and potassium salts of racemic 3-hydroxybutyric acid used in the following examples.

[0052] This article involves the addition amount, content and concentration of various substances, and the percentages mentioned here refer to the mass percentages unless otherwise specified.

[0053] In the embodiments herein, if no specific description is given for the operating temperature, the temperature generally refers to room temperature (15-35° C.).

[0054] HPLC detection method of the components in 3-hydroxybutyric acid betaine salt in the embodiment:

[0055] Instrument: Liquid chromatograph Agilent1260;

[0056] Chromatographic column: Agilent ZORBAX SB-Aq 4.6mm×250mm×5μm or equivalent;

[0057] Mobile phase: 0.1% perchloric acid aqueous solution: acetonitrile = 95:5 (v / v);

[0058] Flow rate: 1.0ml / min;

[0059] Column temperature: 30°C,

[0060] Detector: differential refractometer, detection wavelength: 214nm;

[0061] Injection volume: 10 μl, component content was calculated by area normalization method.

[0062] Among them, the retention time of betaine is 2.599min, the retention time of 3-hydroxybutyric acid is 4.864min, and the retention time of crotonic acid is 15.181min.

[0063] Example 1: Preparation of 3-hydroxybutyric acid betaine sodium salt

[0064]

[0065] The process of forming a salt of sodium 3-hydroxybutyrate and betaine comprises the following steps:

[0066] 1. Weigh 25.00 g (0.198 mol) of sodium 3-hydroxybutyrate and 23.23 g (0.198 mol) of betaine, add them into a three-necked flask, then add 100 ml of deionized water and stir.

[0067] 2. Start the electric heater and slowly raise the temperature to 55°C.

[0068] 3. After the system in the flask is dissolved, continue stirring and reacting at 55℃-60℃ for 3 hours.

[0069] 4. Filter while hot to remove insoluble impurities in the system.

[0070] 5. Start the vacuum pump and concentrate the reaction system under reduced pressure until it becomes viscous, then stop concentrating.

[0071] 6. Slowly add 150 ml of acetone, and white crystals begin to precipitate.

[0072] 7. Cool down to below 10°C and stir for 1 hour.

[0073] 8. Filter and dry the wet product in an oven at 80°C for 4 hours to obtain 47.1 g of 3-hydroxybutyric acid betaine sodium salt as white crystals with a yield of 97.8%.

[0074] 9. The purity of the product and crotonate was determined by HPLC, see Table 1, and the chromatogram is shown in Figure 2 .

[0075] Table 1. Purity of each component in 3-hydroxybutyric acid betaine sodium salt determined by HPLC

[0076] 3-Hydroxybutyric Acid 42.123％ Betaine 57.828％ Crotonic acid 0.049％

[0077] Example 2: Preparation of 3-hydroxybutyric acid betaine potassium salt

[0078] The preparation process of 3-hydroxybutyric acid betaine potassium salt comprises the following steps:

[0079] 1. Weigh 28.17 g (0.198 mol) of potassium 3-hydroxybutyrate and 23.23 g (0.198 mol) of betaine, add them into a three-necked flask, then add 120 ml of deionized water and stir.

[0080] 2. Start the electric heater and slowly raise the temperature to 55°C.

[0081] 3. After the system in the flask is dissolved, continue stirring and reacting at 55℃-60℃ for 3 hours.

[0082] 4. Filter while hot to remove insoluble impurities in the system.

[0083] 5. Start the vacuum pump and concentrate the reaction system under reduced pressure until it becomes viscous, then stop concentrating.

[0084] 6. Slowly add 150 ml of acetone, and white crystals begin to precipitate.

[0085] 7. Cool down to below 10°C and stir for 1 hour.

[0086] 8. Filter and dry the wet product in an oven at 80°C for 4 hours to obtain 48.8 g of 3-hydroxybutyric acid betaine potassium salt as white crystals with a yield of 95%.

[0087] 9. The purity of the product and crotonate was determined by HPLC, see Table 2.

[0088] Table 2. Purity of each component in 3-hydroxybutyric acid betaine potassium salt determined by HPLC

[0089]

[0090]

[0091] Example 3: Preparation of 3-hydroxybutyric acid betaine magnesium salt

[0092] The preparation process of 3-hydroxybutyric acid betaine magnesium salt comprises the following steps:

[0093] 1. Weigh 22.9 g (0.099 mol) of magnesium 3-hydroxybutyrate and 23.23 g (0.198 mol) of betaine, add them into a three-necked flask, then add 250 ml of deionized water and stir.

[0094] 2. Start the electric heater and slowly raise the temperature to 55°C.

[0095] 3. After the system in the flask is dissolved, continue stirring and reacting at 55℃-60℃ for 3 hours.

[0096] 4. Filter while hot to remove insoluble impurities in the system.

[0097] 5. Start the vacuum pump and concentrate the reaction system under reduced pressure until it becomes turbid, then stop concentrating.

[0098] 6. Slowly add 100 ml of ethyl acetate, at which time a large amount of white crystals precipitate.

[0099] 7. Cool down to below 10°C and stir for 1 hour.

[0100] 8. Filter and dry the wet product in an oven at 80°C for 4 hours to obtain 44.2 g of 3-hydroxybutyric acid betaine magnesium salt as white crystals with a yield of 96%.

[0101] 9. The purity of the product and crotonate was determined by HPLC, see Table 3.

[0102] Table 3. Purity of each component in 3-hydroxybutyric acid betaine magnesium salt determined by HPLC

[0103] 3-Hydroxybutyric Acid 42.432％ Betaine 57.385％ Crotonic acid 0.183％

[0104] Example 4: Preparation of 3-hydroxybutyric acid betaine calcium salt

[0105] The preparation process of 3-hydroxybutyric acid betaine calcium salt comprises the following steps:

[0106] 1. Weigh 24.4 g (0.099 mol) of calcium 3-hydroxybutyrate and 23.23 g (0.198 mol) of betaine, add them into a three-necked flask, then add 380 ml of deionized water and stir.

[0107] 2. Start the electric heater and slowly raise the temperature to 55°C.

[0108] 3. After the system in the flask is dissolved, continue stirring and reacting at 55℃-60℃ for 3 hours.

[0109] 4. Filter while hot to remove insoluble impurities in the system.

[0110] 5. Start the vacuum pump and concentrate the reaction system under reduced pressure until a large amount of crystals appear, then stop concentrating.

[0111] 6. Cool down to below 10°C and stir for 1 hour.

[0112] 7. Filter and dry the wet product in an oven at 80°C for 4 hours to obtain 43.1 g of 3-hydroxybutyric acid betaine calcium salt as white crystals with a yield of 91%.

[0113] 9. The purity of the product and crotonate was determined by HPLC, see Table 4.

[0114] Table 4. Purity of each component in 3-hydroxybutyric acid betaine calcium salt determined by HPLC

[0115] 3-Hydroxybutyric Acid 42.143％ Betaine 57.815％ Crotonic acid 0042％

[0116] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them. For those skilled in the art, the present invention may have various modifications and variations. 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 3-hydroxybutyric acid betaine salt having the following molecular structure: in, n is a natural number 1 or 2; M is an alkali metal or an alkaline earth metal selected from Na, K, Mg and Ca; that is, M n+ is a metal ion selected from Na + , K + Mg 2+ and Ca 2+ ;n + is the charge number, representing 1-2 positive charges, The 3-hydroxybutyric acid is selected from D-3-hydroxybutyric acid, L-3-hydroxybutyric acid and DL-3-hydroxybutyric acid.

2. The 3-hydroxybutyric acid betaine salt according to claim 1, characterized in that The equivalent ratio of betaine to 3-hydroxybutyrate is 1:0.3-3.

3. The 3-hydroxybutyric acid betaine salt according to claim 1, characterized in that The equivalent ratio of betaine to 3-hydroxybutyrate is 1:

1.

4. The 3-hydroxybutyric acid betaine salt according to claim 1, characterized in that The crotonic acid content is not higher than 0.2 wt%.

5. A method for preparing the 3-hydroxybutyric acid betaine salt according to any one of claims 1 to 4, characterized in that: When the salt is a sodium salt, a potassium salt or a magnesium salt, the method comprises the following steps: (1) dissolving betaine and 3-hydroxybutyric acid metal salt in a predetermined equivalent ratio in water to obtain an aqueous solution; (2) heating and stirring the aqueous solution until the system is dissolved and clear, and continuing to stir and react for 1-5 hours; (3) Filter while hot to remove insoluble impurities in the system; (4) concentrating the filtrate in step (3) to remove water and form a composite salt; (5) adding an organic solvent to the composite salt obtained in step (4) until crystals are precipitated; (6) standing and cooling until the crystals are almost completely precipitated; (7) filtering and drying to obtain 3-hydroxybutyric acid betaine salt, and When the salt is a calcium salt, the method comprises the following steps: (1) dissolving betaine and calcium 3-hydroxybutyrate in a predetermined equivalent ratio in water to obtain an aqueous solution; (2) heating and stirring the aqueous solution until the system is dissolved and clear, and continuing to stir and react for 1-5 hours; (3) Filter while hot to remove insoluble impurities in the system; (4) concentrating the filtrate in step (3) until crystals are precipitated; (6) standing and cooling until the crystals are almost completely precipitated; (7) filtering and drying to obtain 3-hydroxybutyric acid betaine calcium salt.

6. The method according to claim 5, characterized in that The organic solvent described in step (5) is selected from: ethanol, isopropanol, isopentanol, acetone, ethyl acetate or a mixture of two or more thereof.

7. The method according to claim 5, characterized in that The stirring reaction temperature in step (2) is below 65°C.

8. The method according to claim 5, characterized in that The method further comprises the following steps: (8) recovering the organic solvent by distillation or rectification.

9. Use of the 3-hydroxybutyric acid betaine salt according to any one of claims 1 to 4 in food and medicine.

10. The use according to claim 9, characterized in that The food is used to reduce or prevent fatty liver or protect kidneys, and the medicine is used to treat fatty liver or protect kidneys.

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