WO2019034112A1 - COMPOSITION CONTAINING L-CARNITINE AND β-HYDROXYBUTYRATE COMPOUND - Google Patents

Abstract

Provided are a composition containing L-carnitine and at least one β-hydroxybutyrate compound; said β-hydroxybutyrate compound contains β-hydroxybutyrate and a β-hydroxybutyrate precursor or a combination thereof; preferably, the L-carnitine and β-hydroxybutyrate in said composition are present in the form of an intramolecular salt. Also provided is a method for preparing L-carnitine-β-hydroxybutyrate. The composition has weight-loss and ketogenic effectiveness.

Description

Composition comprising L-carnitine and β-hydroxybutyric acid compound Technical field

The invention belongs to the technical field of medicine, and particularly relates to a composition comprising L-carnitine and β-hydroxybutyric acid compound, and the invention also relates to preparation of L-carnitine-β-hydroxybutyrate in medicine and food additive Application in technology.

Background technique

Obesity has become a serious public health problem. In recent years, obesity patients are increasing all over the world. The number of deaths due to obesity is second only to smoking every year. The International Obesity Conference released a report on the number of deaths caused by obesity worldwide. The number of people who have starved to death in the world has been exceeded. Obesity can also cause severe diabetes, cardiovascular disease and other diseases, and is the enemy of human health and longevity.

Therapeutic drugs for obesity include: appetite suppressing drugs, drugs for increasing energy consumption, drugs for inhibiting intestinal digestion and absorption, insulin sensitizers, biological peptides, and agonists or inhibitors thereof. These drugs have many adverse reactions, such as heart valve damage, pulmonary hypertension and finger necrosis, elevated blood pressure, gastrointestinal dysfunction, respiratory infections, headache, menstrual disorders, anxiety, fatigue, flatulence, abdominal pain, diarrhea, etc. Individual patients may also have hypoglycemia and hepatotoxicity.

The main physiological function of L-carnitine is to promote the conversion of fat into energy. L-carnitine can reduce body fat and reduce body weight without reducing water and muscle. It was recognized as the safest and no side effect by the International Obesity Health Organization in 2003. Weight loss nutritional supplements. L-carnitine is a key substance in the process of fat metabolism, which can promote the oxidative decomposition of fatty acids into mitochondria. L-carnitine is a carrier for transporting fatty acids. In long-term high-intensity exercise, L-carnitine increases the rate of fat oxidation, reduces the consumption of glycogen, and also delays fatigue. Many weight loss products use L-carnitine as one of the main components of weight loss.

L-carnitine has been used in the fields of medicine, health care and food, and has been regulated as a legal multi-purpose nutrient by Switzerland, France, the United States and the World Health Organization. China's food additive health standard GB2760-1996 stipulates that L-carnitine tartrate is a food nutrition fortifier, which can be applied to chewable tablets, drinking liquids, capsules, milk powder and milk drinks. In addition, medical effects including cardiovascular disease, liver disease, kidney disease, hyperlipidemia, diabetes, neuromuscular diseases, etc. can be improved by taking L-carnitine and its products.

It is also known that the pharmaceutically acceptable salt of L-carnitine has the same therapeutic or nutritional effect as its internal salt. And these pharmaceutically acceptable salts can improve the stability and moisture absorption of the inner salt, such as L-carnitine tartrate (US4602039) and L-fumarate (US2009281183A1), L-carnitine mucate commonly used in the market. (US5952379). It has also been reported that L-carnitine is combined with metal ions such as calcium and magnesium ions to form a salt which has good water solubility and is easily absorbed by the human body. For example, US Pat. No. 6,051,608 discloses magnesium levoth fumarate and alkanoyl L-carnitine magnesium fumarate, WO 98/45250 discloses L-carnitine magnesium tartrate and alkanoyl L-carnitine magnesium citrate; there are also patent reports of L-carnitine calcium galactate and L-carnitine calcium fumarate (WO2008080287).

However, the existing pharmaceutically acceptable salts of L-carnitine still have some problems and defects, such as L-carnitine tartrate still has a large moisture absorption, relative humidity above 60% will deliquesce; and L-carnitine and metal ions such as Salts formed by the combination of calcium and magnesium ions also have some defects. For example, a large amount of calcium lactate may cause fatigue; excessive calcium gluconate and calcium chloride are unfavorable for diabetes; in addition, L-carnitine calcium galactate is in the preparation process. There is also a risk of insufficient supply of raw materials, and the cost is high, because lactobionic acid is not produced in large quantities in China, and the price is relatively expensive; the process of preparation of L-carnitine calcium fumarate (CN101209975) needs to be frozen, which is in the process of large production. The operation is inconvenient and the production cost is increased.

Studies have also shown that the pursuit of weight loss through the ketogenic diet may result in loss of fat storage while maintaining and protecting muscle mass; some studies have also shown that the muscle-protective nature of the ketogenic diet leads to an increase in physical performance.

Nutritional or therapeutic ketosis is a physiological condition caused by a ketogenic diet, calorie restriction, therapeutic fasting, and/or supplementation with ketogenic precursors resulting in elevated blood ketone levels (typically above 0.5 mmol/L). Ketone bodies represent alternative energy substrates for peripheral tissues and the central nervous system. Beta-hydroxybutyrate and acetoacetate are the two most abundant and physiologically significant ketone bodies. Glucose storage in the body is quickly used and becomes rapidly depleted during fasting, extreme sports, and/or low carbohydrate consumption. Failure to replenish glucose storage as they become depleted causes the body to resort to alternative methods to produce energy by making ketone bodies. Ketone bodies can be used as an alternative fuel by every cell in the body to meet the body's energy needs, including the needs of the brain. During delayed fasting, for example, blood ketone levels will increase up to 2 mmol/L or 3 mmol/L. It is conventionally understood and agreed that when the blood ketone rises above 0.5 mmol/L, the heart, brain and peripheral tissues use ketone bodies (β-hydroxybutyric acid and acetoacetic acid) as the main fuel source. This state is called Ketoacidosis or "nutritional ketosis". When in ketosis, the body burns fat essentially, so once in ketosis, one can induce body fat loss by reducing dietary fat intake and adjusting carbohydrate intake to a low enough to maintain ketosis.

Although the pursuit of weight loss through the ketogenic diet has many advantages, and then, in the pursuit and maintenance of the ketone lifestyle, there are significant obstacles, such as the difficulty of transition to the ketogenic state, most people are limited by the ability to enter ketosis The fastest way to deplete glucose in the body is through fasting and exercise, which is physically and emotionally demanding for those who get pleasure from eating cakes, candy, bread, etc. And it has great challenges; when entering the transition to ketosis, there will be uncomfortable physical and mental states such as drowsiness and dizziness.

Although it has been reported that ketogenic medical foods or exogenous ketones can contribute to the transition to ketosis, ketogenic fats such as medium chain triglycerides (MCT oils) are usually not required by the gastrointestinal system to help induce ketosis. The amount is well tolerated; and the oral administration of beta-hydroxybutyrate (βHB) and acetoacetate in its free acid form is expensive and ineffective in the manufacture of persistent ketosis. The sodium salt is used to buffer the free acid form of [beta]HB, but this causes potentially harmful sodium overload and mineral balance imbalance at the therapeutic level of ketosis. It has also been suggested that the ketogenic diet raises concerns about increasing total cholesterol and triglycerides while lowering high-density lipoprotein (HDL) levels, leading to cardiovascular disease.

Summary of the invention

The object of the present invention is to overcome the above-mentioned deficiencies of the prior art, to provide a novel composition comprising L-carnitine; and at least one β-hydroxybutyric acid compound comprising β-hydroxyl groups Butyrate, beta-hydroxybutyrate precursor or a combination thereof.

Another aspect of the present invention also provides a composition comprising L-carnitine and a β-hydroxybutyric acid compound in the form of an intramolecular salt.

A third aspect of the invention provides a process for the preparation of the composition and the use of the composition.

Preferably, the present invention provides a composition comprising L-carnitine; and at least one β-hydroxybutyric acid compound comprising β-hydroxybutyrate, β-hydroxybutyric acid Precursor or a combination thereof.

Wherein the at least one β-hydroxybutyric acid compound comprises one or more of the following: sodium β-hydroxybutyrate, potassium β-hydroxybutyrate, calcium β-hydroxybutyrate, magnesium β-hydroxybutyrate , lithium beta-hydroxybutyrate, or a mixture thereof;

--hydroxybutyrate arginine, β-hydroxybutyrate lysine, β-hydroxybutyrate histidine, β-hydroxybutyrate ornithine, β-hydroxybutyrate sarcosine, β-hydroxybutyric acid Mercaptoamine or β-hydroxybutyric acid citrulline;

1,3-butanediol, ethyl acetoacetate or β-hydroxybutyrate; or

a combination of β-hydroxybutyrate with 1,3-butylene glycol, ethyl acetoacetate or β-hydroxybutyrate; or

A combination of a β-hydroxybutyrate mixture with 1,3-butanediol, ethyl acetoacetate or β-hydroxybutyrate.

Further preferably, the at least one β-hydroxybutyric acid compound comprises

(1) sodium β-hydroxybutyrate, β-hydroxybutyrate arginine or a combination thereof; or

(2) a combination of a sodium salt of β-hydroxybutyrate and a potassium salt of β-hydroxybutyrate; or

(3) β-hydroxybutyrate histidine, β-hydroxybutyrate ornithine, β-hydroxybutyrate sarcosine, β-hydroxybutyrate decyl butylamine or β-hydroxybutyrate citrulline.

Wherein the at least one β-hydroxybutyric acid compound is racemic DL-β-hydroxybutyric acid or the single isomer R-β-hydroxybutyric acid.

Preferably, in the above composition, the molar ratio of the β-hydroxybutyric acid compound to the L-carnitine is from 20:1 to 1:1.

Further, the present invention provides a composition comprising the compound A _m B _n wherein A is L-carnitine, B is β-hydroxybutyrate arginine, β-hydroxybutyrate lysine, β-hydroxyl Butyric acid histidine, β-hydroxybutyrate ornithine, β-hydroxybutyrate sarcosine, β-hydroxybutyrate decyl butylamine or β-hydroxybutyrate citrulline; m, n independently selected from An integer of from 1 to 4; preferably B is β-hydroxybutyrate arginine, β-hydroxybutyrate lysine, β-hydroxybutyrate histidine, or β-hydroxybutyrate ornithine, and preferably m is 1 or 2, and n is 2 or 1.

In another preferred embodiment of the present invention, there is provided a composition comprising L-carnitine-β-hydroxybutyrate, and wherein the L-carnitine and the β-hydroxybutyric acid compound are intramolecular salts. The form exists and has the following structure:

Wherein M is Na, K or Li, and the ratio of m to n is from 0.8:10 to 10:0.8, preferably from 1:4 to 4:1, more preferably 1:1, most preferably m and n are both 1, or ,

其中M为Ca或Mg，m与n的取值比为0.8:10～10:0.8，优选1:4～4:1，更优选1:1，最优选m与n均为1。

Wherein M is Ca or Mg, and the ratio of m to n is from 0.8:10 to 10:0.8, preferably from 1:4 to 4:1, more preferably 1:1, and most preferably m and n are both 1.

Further, preferably, the composition comprises L-carnitine-β-hydroxybutyrate, and the L-carnitine-β-hydroxybutyrate is a combination of one or more of the following:

Wherein, the ratio of values of m to n is from 0.8:10 to 10:0.8, preferably from 1:4 to 4:1, more preferably 1:1, and most preferably m and n are both 1.

In another preferred embodiment of the present invention, there is provided a composition comprising L-carnitine-β-hydroxybutyrate, the L-carnitine-β-hydroxybutyrate being the following compound ( a mixture of Ia) and compound (Ib),

其中m与n的取值比1:4～4:1，优选1:1；更优选所述化合物Ia和Ib中m，n均为1。

Wherein the ratio of m to n is 1:4 to 4:1, preferably 1:1; more preferably, m and n are 1 in the compounds Ia and Ib.

In another preferred embodiment of the invention, there is provided a composition comprising one or more compounds A _m B _n and/or compound I and/or compound II. Preferably, for example, the composition comprises one or more compounds A _m B _n and one or more compounds I; the composition comprising one or more compounds A _m B _n and one or more Compound II; the composition comprising one or more compounds A _m B _n and one or more compounds I and one or more compounds II;

Wherein compound I is:

其中M为Na,K或Li，m与n的取值比为0.8:10～10:0.8，优选1:4～4:1，更优选m与n均为1；

Wherein M is Na, K or Li, and the ratio of m to n is from 0.8:10 to 10:0.8, preferably from 1:4 to 4:1, more preferably both m and n are 1;

Compound II is:

其中M为Ca或Mg，m与n的取值比为0.8:10～10:0.8，优选1:4～4:1，更优选m与n均为1。

Wherein M is Ca or Mg, and the ratio of m to n is from 0.8:10 to 10:0.8, preferably from 1:4 to 4:1, more preferably both m and n are 1.

Also provided in a second aspect of the invention is the use of the above composition for the preparation of a medicament for the promotion of ketosis in a mammal, preferably a L-carnitine, and at least one of the compositions The β-hydroxybutyric acid compound is used as an active ingredient, or the compound A _m B _n and/or the compound I and/or the compound II are used as an active ingredient.

The above composition provided by the present invention may also be used for the preparation of a slimming or health food, a diet/nutrition supplement, a veterinary product or a feed, preferably, the composition has L-carnitine, and at least one β-hydroxyl group. A butyric acid compound is used as an active ingredient, or a compound A _m B _n and/or a compound I and/or a compound II as an active ingredient.

Preliminary studies have found that the compositions provided by the present invention are also advantageous for treating hyperglycemia or type II diabetes and improving the general health of the user in a short period of time. It can also be used as a brain-enhancing medicine to improve athlete performance, to prevent diseases associated with metabolic disorders, mitochondrial defects and insulin resistance, as an adjunct to the ketogenic diet, as an anti-aging supplement, and for Other uses associated with increased metabolic health.

When the above composition of the present invention is applied, it can be formulated into an oral dosage form including: a tablet, a capsule, a granule, a pill, and an oral solution. The composition further comprises one or more pharmaceutically acceptable or edible excipients, which are formed into a powder mixture, as a ready-to-drink liquid, into hard or soft capsules, tablets, concentrated gels or in the field. Any other suitable dosage form known to the skilled person.

When employed, the compositions described herein and optional pharmacologically acceptable excipients can be combined. According to preliminary calculations, as a slimming drug or a health food, the composition is equivalent to 20-2000 mg/day, or 50 mg to 1000 mg, or 40-800 mg/day, per unit weight of L-carnitine. As a dietary/nutritional supplement, the composition comprises, by weight, L-carnitine in a unit dose equivalent to 50-3000 mg/kg, or 100-1000 mg/kg; as a drug for promoting ketosis, the composition is by weight The β-hydroxybutyric acid in a unit dose corresponds to 1 g to 50 g, or 2 g to 30 g, or 2 g to 10 g, or 5 g to 8 g. The frequency of use and the specific amount of the preferred preparation may vary depending on various factors such as the degree of susceptibility of the individual, the age, sex and weight of the individual, the specific reaction of the individual, etc., and optimization of these factors can be determined by those skilled in the art as needed. Another aspect of the present invention provides a method for preparing the compound A _m B _n , the compound I, and the compound II in the composition;

A _m B _n can be prepared by the following method, wherein m and n have a value of 1:4 to 4:1, preferably 1:1.

Adding a small amount of water to an organic solvent such as acetone, isopropanol or the like as a solvent, and reacting the β-hydroxybutyric acid amino acid salt with L-carnitine (preferably in a molar ratio of 1 to 1.2:1), more preferably, first to rotate left The carnitine is dissolved in a small amount of pure water, and then an equimolar amount of a base such as sodium hydroxide or potassium hydroxide is slowly added dropwise, and then the prepared L-carnitine solution is added to the above-mentioned organic compound of the β-hydroxybutyric acid amino acid salt. The reaction is carried out in a solvent. After completion of the reaction, the solvent was removed under reduced pressure in a water bath at 40 ° C, and an organic solvent such as acetone, isopropyl alcohol or the like was added to the flask, and the mixture was stirred to precipitate a solid, which was filtered and dried in vacuo to give a product.

Compound I and Compound II can be prepared as follows:

Dissolving 3-hydroxybutyric acid in an organic solvent such as acetone, isopropanol or dichloromethane to obtain a solution of 3-hydroxybutyric acid; further dissolving L-carnitine in pure water and adding a base to the solution ( An aqueous solution such as sodium hydroxide, potassium hydroxide, lithium hydroxide, calcium hydroxide or magnesium hydroxide. The L-carnitine solution is slowly dropped into a solution of 3-hydroxybutyric acid, and then the mixed reaction solution is removed under reduced pressure in a water bath at 35 ° C to 45 ° C, preferably 40 ° C, and then left to the residue. Add the corresponding organic solvent (such as acetone, isopropanol or dichloromethane), stir and crystallize, filter, filter cake washed with organic solvent (such as acetone, isopropanol or dichloromethane) and vacuum dried to obtain the corresponding compound I , Compound II. The organic solvent is preferably acetone; the base used is selected according to the corresponding metal in the compound I or the compound II. For example, when M in the compound I is Na, the base used is preferably sodium hydroxide. The present invention provides L-carnitine; and at least one β-hydroxybutyric acid compound comprising β-hydroxybutyrate, β-hydroxybutyrate precursor or a combination thereof The composition is capable of significantly promoting fat metabolism, and synergistically promotes a fat loss effect by significantly increasing blood ketone and lowering blood sugar level, and the use of the composition does not adversely affect the lipid mass spectrum, by using the composition, The rapid initiation of ketosis and the rate at which ketone adaptation is accelerated is beneficial to avoid the symptoms of glucose withdrawal that are routinely experienced by individuals who initiate a ketogenic diet. It is more easily accepted by the human body than the existing L-carnitine salt, and is more likely to participate in physiological metabolism of the human body, and has stronger nutrition and therapeutic effects.

These compounds in the form of intramolecular salts in the compositions comprising one or more of the compounds A _m B _n and/or Compound I and/or Compound II provided by the present invention have a more significant advantage:

First, when the human body takes these compositions containing L-carnitine and β-hydroxybutyric acid compounds, the compound A _m B _n , the β-hydroxybutyric acid precursor or the β-hydroxybutyrate in the compound I and the compound II structure. Part of it will not accumulate in the human body, but can be used as a ketone supplement to enable patients to quickly establish ketosis and maintain ketosis with little or no perception of the patient's physical and psychological comfort; The ketone body also did not find a decrease in high-density lipoprotein (HDL) levels.

Secondly, after A _m B _n , Compound I, and Compound II enter the body, the L-carnitine part promotes the decomposition of the fat while promoting the ketone body supplement (such as β-hydroxybutyric acid compound, β-hydroxybutyrate or Oxidation of β-hydroxybutyric acid precursors, thereby promoting the utilization and elimination of these ketone bodies, reducing metabolic toxicity caused by acyl accumulation during ketone body utilization; in particular, A _m B _n , Compound I In the compound II, the values of m and n are 1:4 to 4:1, especially when m and n are simultaneously 1, the ketogenic and weight loss effects are the best;

Thirdly, in the pharmaceutical composition, L-carnitine and β-hydroxybutyric acid compound are present in the form of intramolecular salt compounds A _m B _n , Compound I and Compound II, which are more stable in nature, are not hygroscopic, and are bioavailable in vivo. Higher degrees.

Fourth, when the composition of the present invention comprises the compound A _m B _n and when B is a β-hydroxybutyric acid amino acid (such as β-hydroxybutyrate arginine, β-hydroxybutyrate lysine, β - when using hydroxybutyrate histidine, beta-hydroxybutyrate ornithine, beta-hydroxybutyrate sarcosine, beta-hydroxybutyrate decylamine or beta-hydroxybutyrate citrulline, etc. At the same time, it also increases the amino acid supplement in the human body, which is very beneficial for users who need to lose weight or ketone.

Fifth, the composition comprising the compound A _m B _n and when B is the above-mentioned β-hydroxybutyric acid amino acid, on the other hand, also solves the possibility that the presence of β-hydroxybutyric acid in the form of a mineral salt may cause potential Mineral salt overload, such as sodium salt overload, the risk of cardiovascular effects of calcium salts.

Sixth, the composition provided by the invention has simple and reasonable production process, and reacts under normal pressure, mild reaction conditions, and is suitable for industrial large-scale production.

Detailed ways

The advantages of the technical solutions of the present invention are specifically explained below in conjunction with the specific embodiments. It is to be understood that the embodiments are merely exemplary and not intended to limit the invention.

The "β-hydroxybutyric acid" and the "3-hydroxybutyrate" described in the present invention are the same compound and are mutually replaceable. The "L-carnitine-β-hydroxybutyrate" refers to an internal salt formed by L-carnitine and a β-hydroxybutyric acid compound; and the “β-hydroxybutyrate” refers to β-hydroxybutyric acid. a metal salt such as a sodium salt, a potassium salt, a calcium salt, a magnesium salt, and a salt of β-hydroxybutyric acid and an amino acid; the “β-hydroxybutyric acid amino acid salt” refers to a β-hydroxybutyric acid and an amino acid. salt.

Example 1, preparation method of compound (Iaa):

To a 100 mL round bottom flask was added 1.04 g (10 mmol) of 3-hydroxybutyric acid and 8 mL of acetone. 1.61 g (10 mmol) of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of a 10 N (10 mmol) aqueous sodium hydroxide solution was added to the solution. The prepared L-carnitine solution was slowly dropped into the reaction flask, and the reaction was carried out for 5 minutes after the dropwise addition. The solvent was removed under reduced pressure in a water bath of 40 ° C to give a colorless oil, and 80 mL of acetone was added to the flask. The crystals were stirred and filtered, and the filter cake was washed with acetone and dried in vacuo to yield 2.58 g of white solid.

Chemical Formula: C ₁₁ H ₂₂ NO ₆ Na; 1H NMR (D ₂ O): δ 1.12 (3H, d), 2.12 to 2.41 (4H, m), 3.14 (9H, s), 3.34 (2H, dd), 4.04 to 4.09 (1H, m), 4.47 to 4.50 (1H, m).

Example 2: Preparation of Compound (IIaa) of the following formula

To a 100 mL round bottom flask was added 1.04 g (10 mmol) of 3-hydroxybutyric acid and 16 mL of acetone. 1.61 g (10 mmol) of L-carnitine was dissolved in 16 mL of pure water, and 0.37 g (5 mmol) of calcium hydroxide powder was added to the solution, and the mixture was heated to 80 ° C for 2 hours, and then the suspension of the L-carnitine solution was dropped. The reaction mixture was heated to 60 ° C and stirred for 2 hours, and then dissolved. The solvent was removed under reduced pressure in a water bath of 40 ° C to obtain a creamy white oil. To the flask was added 80 mL of acetone, stirred and crystallized, filtered, and filtered cake. After washing with acetone and vacuum drying, 1.76 g of a white product was obtained, yield 86%.

_{Chemical Formula: C 22 H 44 N} 2 O 12 Ca; 1 HNMR (D 2 O): δ1.13 (3H, d), 2.11 ~ 2.40 (8H, m), 3.15 (18H, s), 3.36 (4H, Dd), 4.05 to 4.13 (2H, m), 4.47 to 4.51 (2H, m).

Example 3: Preparation of L-carnitine-β-hydroxybutyrate arginine (in which the molar ratio of L-carnitine to β-hydroxybutyrate arginine is 1:1):

To a 100 mL round bottom flask was added 2.78 g (11 mmol) of β-hydroxybutyrate arginine, and 10 mL of acetone and 3 mL of purified water were added. 1.61 g (10 mmol) of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of 10 N (10 mmol) sodium hydroxide solution was added to the solution. The L-carnitine solution was slowly dropped into the reaction flask. After the completion of the dropwise addition, the reaction was carried out for 5 minutes. The solvent was removed under reduced pressure in a water bath of 40 ° C to give a colorless oil, and 80 mL of acetone was added to the flask. After stirring for 20 minutes, a white solid was precipitated, filtered, and the filter cake was washed with acetone and dried in vacuo to give 3.92 g of white product.

Example 4: Preparation of L-carnitine-β-hydroxybutyrate lysine (in which the molar ratio of L-carnitine to β-hydroxybutyrate lysine is 1:1):

To a 100 mL round bottom flask was added 11 mmol of β-hydroxybutyrate lysine, and 10 mL of acetone and 3 mL of purified water were added. 10 mmol of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of 10 N (10 mmol) sodium hydroxide solution was added to the solution. The L-carnitine solution was slowly dropped into the reaction flask. After the completion of the dropwise addition, the reaction was carried out for 5 minutes. The solvent was removed under reduced pressure in a water bath of 40 ° C to give a colorless oil, and 80 mL of acetone was added to the flask. After stirring for 20 minutes, a white solid was precipitated and filtered, and the filter cake was washed with acetone and dried in vacuo to give 3.92 g of white product.

Example 5: Preparation of L-carnitine-β-hydroxybutyrate ornithine (in which the molar ratio of L-carnitine to β-hydroxybutyrate ornithine is 1:1):

To a 100 mL round bottom flask was added 11 mmol of β-hydroxybutyrate ornithine, and 10 mL of acetone and 3 mL of purified water were added. 10 mmol of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of 10 N (10 mmol) sodium hydroxide solution was added to the solution. The L-carnitine solution was slowly dropped into the reaction flask. After the completion of the dropwise addition, the reaction was carried out for 5 minutes. The solvent was removed under reduced pressure in a water bath of 40 ° C to give a colorless oil, and 80 mL of acetone was added to the flask. After stirring for 20 minutes, a white solid was precipitated, filtered, and the filter cake was washed with acetone and dried in vacuo to give 3.92 g of white product.

Example 6: Preparation of L-carnitine-β-hydroxybutyrate amdecylamine (in which the molar ratio of L-carnitine to β-hydroxybutyrate is 2:1)

To a 100 mL round bottom flask was added 10.5 mmol of β-hydroxybutyric acid decylamine, and 10 mL of acetone and 3 mL of purified water were added. 10 mmol of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of 10 N (10 mmol) sodium hydroxide solution was added to the solution. The prepared L-carnitine solution was slowly dropped into the reaction flask. After the completion of the dropwise addition, the reaction was carried out for 5 minutes. The solvent was removed under reduced pressure in a water bath at 40 ° C, and 80 mL of acetone was added to the flask, and the mixture was stirred for 20 minutes to precipitate white. The solid was filtered, and the filter cake was washed with acetone and dried in vacuo to give 3.92 g of white product.

Example 8: Preparation of L-carnitine-β-hydroxybutyrate histidine (in which the molar ratio of L-carnitine to β-hydroxybutyrate histidine is 1:2)

To a 100 mL round bottom flask was added 11 mmol of β-hydroxybutyrate histidine, and 10 mL of acetone and 3 mL of purified water were added. 10 mmol of L-carnitine was dissolved in 3 mL of pure water, and 1 mL of 10 N (10 mmol) sodium hydroxide solution was added to the solution. The prepared L-carnitine solution was slowly dropped into the reaction flask. After the completion of the dropwise addition, the reaction was carried out for 5 minutes. The solvent was removed under reduced pressure in a water bath at 40 ° C, and 80 mL of acetone was added to the flask, and the mixture was stirred for 20 minutes to precipitate white. The solid was filtered, and the filter cake was washed with acetone and dried in vacuo to give 3.92 g of white product.

Example 9: Study on weight loss effect, ketogenic effect and lipid mass spectrometry:

56 SPF male SD rats were purchased from Shanghai Slack Laboratory Animals Co., Ltd. The experimental animals were kept in the SPF animal room. The animal room was well ventilated and equipped with air conditioning. The temperature was kept at 20-25 °C, the humidity was kept at 40%-70%, the air exchange times were 10-15 times/h, and the light and dark lighting was 12 hours each. Experimental animals were given free access to food and water, and each rat was labeled with an ear tag. Eight blank rats were fed normal diet, and 48 model rats were fed a high-fat and high-cholesterol diet (Nantong Trophy Feed Technology Co., Ltd.) for 60 days to establish an obesity model. According to the body weight, 48 rats were randomly divided into six groups: model group, model + exercise group, composition (compound prepared in Example 1 (Iaa)) + exercise group, physical mixing (L-carnitine and β- Sodium hydroxybutyrate molar ratio 1:1 mixed) + exercise group, L-carnitine + exercise group and β-hydroxybutyrate + exercise group, 8 / group. The blank control group and the model group were only routinely reared, and there was no other intervention. The rats in the exercise group were forced to exercise for half an hour every day. Rats in the blank control group, model group and model + exercise group were given normal saline, 10 mL/kg once a day for 4 weeks. Composition (Compound (Iaa) prepared in Example 1) + exercise group, physical mixing (mixed L-carnitine with β-hydroxybutyrate molar ratio 1:1) + exercise group, L-carnitine + exercise group and β- The hydroxybutyrate + exercise group was intragastrically administered with 15 mmol/kg of the composition, physical mixture, L-carnitine and β-hydroxybutyric acid once a day for 4 weeks. The body weight of the animals was measured four weeks after the administration, and the ketone body level was measured using a Yicheng T-1 blood ketone body tester 4 hours after the last administration, and serum triglyceride (TG) and cholesterol (TCHO) were determined by blood separation serum. , low density lipoprotein (LDL) and high density lipoprotein (HDL) levels. The experimental results are shown in Table 1 below.

Table 1: Results of weight loss, ketogenic effects and lipid mass spectrometry:

L-carnitine-β-hydroxybutyrate arginine prepared in Example 3, L-carnitine-β-hydroxybutyrate ornithine prepared in Example 5 was substituted for the above compound (Iaa), and the corresponding L-carnitine was used. a 1:1 physical mixture with a molar ratio of β-hydroxybutyrate arginine, a 1:1 physical mixture of L-carnitine and β-hydroxybutyrate ornithine molar ratio in place of the above physical mixture, and the same operation was carried out. A similar trend; L-carnitine-β-hydroxybutyrate arginine or L-carnitine-β-hydroxybutyrate ornithine + exercise group had the best weight loss effect, and the ketogenic effect was the most significant and increased to some extent. HDL; L-carnitine and β-hydroxybutyrate (such as β-hydroxybutyrate ornithine, β-hydroxybutyrate arginine) + exercise group weight loss and ketone effect is also significantly higher than the use of left-handed meat alone In the base + exercise group or the β-hydroxybutyrate + exercise group, it was found that the β-hydroxybutyrate + exercise group reduced HDL to a certain extent.

Example 10: In vivo bioavailability study

Twelve SPF male SD rats were purchased from Shanghai Slack Laboratory Animals Co., Ltd. The experimental animals were kept in the SPF animal room. The animal room was well ventilated and equipped with air conditioning. The temperature was kept at 20-25 °C, the humidity was kept at 40%-70%, the air exchange times were 10-15 times/h, and the light and dark lighting was 12 hours each. Experimental animals were given free access to food and water, and each rat was labeled with an ear tag. Animals before the administration were fasted overnight, and were randomly divided into 4 groups and 3 groups/group according to body weight. Three rats were given 1 mg/kg L-carnitine intravenously, 3 rats were given 5 mg/kg L-carnitine-β-hydroxybutyrate, and 3 rats were given 5 mg/kg L-carnitine and β-. Physical mixture of hydroxybutyrate, 3 rats were intragastrically administered with 5 mg/kg L-carnitine, before the administration and 5, 15, 30, 1, 2, 4, 8, 12 and 24 h after the completion of the bolus, by the neck 0.2 mL of venous blood was collected and placed in a test tube containing K2-EDTA anticoagulant. The whole blood sample collected was placed on ice before centrifugation. The whole blood sample collected at the same time point was centrifuged within half an hour after the collection was completed. After centrifugation (5500 rpm) for 10 min, the plasma was separated and stored in a -70 ° C refrigerator. The concentration of L-carnitine in rat plasma was determined by LC-MS/MS analysis. The pharmacokinetic parameters were calculated using the WinNonlin non-compartment model, and the absolute bioavailability was calculated. The results are shown in Table 2.

Table 2: Bioavailability

The L-carnitine-β-hydroxybutyrate prepared in Examples 3 to 8 of the present invention was physically mixed with L-carnitine and its corresponding β-hydroxybutyrate to carry out the above bioavailability study, and the experimental results showed similar to Table 2 The trend is that L-carnitine-β-hydroxybutyrate has the highest bioavailability, followed by a physical mixture, which is more bioavailable than L-carnitine alone.

Example 11: Stability study

The compound (Iab), the compound (IIaa), and the L-carnitine-β-hydroxybutyric acid prepared in Examples 3 to 8 were tested according to the Guiding Principles of Drug Hygroscopicity (Chinese Pharmacopoeia 2015 Edition, Part 4, 9103). Salt, and the corresponding physical mixture of L-carnitine and β-hydroxybutyrate, and L-carnitine-calcium fumarate, the hygroscopicity results are shown in Table 3.

Table 3: Humidity results

product Humidity Compound (Iaa) 0.1% Compound (IIaa) 0.2% L-carnitine-β-hydroxybutyrate <0.3% Physical mixture <0.5% L-carnitine-calcium fumarate 0.8%

Claims (11)

A composition comprising L-carnitine, and at least one beta-hydroxybutyrate compound comprising beta-hydroxybutyrate, beta-hydroxybutyrate precursor or a combination thereof. The composition according to claim 1, wherein said at least one β-hydroxybutyric acid compound comprises one or more of the following: sodium β-hydroxybutyrate, potassium β-hydroxybutyrate, β- Calcium hydroxybutyrate, magnesium β-hydroxybutyrate, lithium β-hydroxybutyrate, or a mixture thereof; β-hydroxybutyrate arginine, β-hydroxybutyrate lysine, β-hydroxybutyrate histidine,鸟-hydroxybutyric acid ornithine, β-hydroxybutyric acid sarcosine, β-hydroxybutyrate decyl butylamine or β-hydroxybutyric acid citrulline; 1,3-butanediol, ethyl acetoacetate or Ethyl β-hydroxybutyrate; or a combination of β-hydroxybutyrate with 1,3-butanediol, ethyl acetoacetate or β-hydroxybutyrate; or a mixture of β-hydroxybutyrate and 1, A combination of 3-butanediol, ethyl acetoacetate or β-hydroxybutyrate. The composition of claim 1 wherein said at least one beta-hydroxybutyric acid compound comprises (1) sodium β-hydroxybutyrate, β-hydroxybutyrate arginine or a combination thereof; or (2) a combination of a sodium salt of β-hydroxybutyrate and a potassium salt of β-hydroxybutyrate; or (3) β-hydroxybutyrate histidine, β-hydroxybutyrate ornithine, β-hydroxybutyrate sarcosine, β-hydroxybutyrate decyl butylamine or β-hydroxybutyrate citrulline. The composition according to claim 1, wherein said at least one β-hydroxybutyric acid compound is racemic DL-β-hydroxybutyric acid or a single isomer R-β- Hydroxybutyric acid. The composition according to any one of claims 1 to 4, wherein the molar ratio of the β-hydroxybutyric acid compound to the L-carnitine in the composition is from 20:1 to 1:1, preferably 1:4. ~4:1. A composition comprising a compound A _m B _n , wherein A is L-carnitine, B is β-hydroxybutyrate arginine, β-hydroxybutyrate lysine, β-hydroxybutyrate histidine, β - hydroxybutyric acid ornithine, β-hydroxybutyric acid sarcosine, β-hydroxybutyrate decyl butylamine or β-hydroxybutyric acid citrulline; m, and n have a value of 1:4 to 4: 1. The composition of claim 6 wherein said m, n are both 1. The composition according to claim 7, further comprising a compound of the formula I or II:

Wherein M is Na, K or Li, and the ratio of m to n is from 1:4 to 4:1, preferably 1:1; or

Wherein M is Ca or Mg, and the ratio of m to n is from 1:4 to 4:1, preferably 1:1. Use of a composition according to any one of claims 1 to 8 for the preparation of a medicament/nutraceutical for promoting ketosis in a mammal. Use of a composition according to any one of claims 1 to 8 for the preparation of a slimming or health food, a diet/nutrition supplement, a veterinary product or a feed. Use of a composition according to any one of claims 9 to 10, characterized in that the composition is formulated into an oral dosage form comprising: a tablet, a capsule, a granule, a pill, an oral solution.