Composition, pharmaceutical preparation, food or health care product and application thereof

Abstract

The invention discloses a composition, a pharmaceutical preparation, a food or a health-care product and application thereof. According to the application, the NAD + depletion and metabolic disorder degree after drinking can be relieved by combining NAM on the basis of a medicine (tetramethylpyrazine) for promoting the NAD + remedial synthesis capability, and liver injury caused by alcohol is expected to be relieved fundamentally. Experiments show that the combination of 50 mg / kg of TMP and 250 mg / kg of NAM for promoting synthesis of NAD + can effectively relieve liver injuries such as alcoholic lipid degeneration and the like.

Description

Technical Field

[0001] The present invention generally relates to the field of biomedical technology, and in particular to a composition, a pharmaceutical preparation, a food or a health product and uses thereof. Background Art

[0002] Alcoholic Liver Disease (ALD) is one of the most common liver diseases in the world, with a high incidence, complex disease stages, and a lack of targeted therapeutic drugs. Currently, the ALD treatment plans in domestic and international guidelines are based on strict abstinence from alcohol. However, due to personal and social factors, strict abstinence from alcohol is often difficult to implement smoothly. Therefore, how to minimize the harm of alcohol to the human body and timely and effectively intervene and treat ALD at an early stage is one of the important issues in the prevention and control of ALD.

[0003] Alcohol metabolism is mainly divided into two stages: first, alcohol molecules are oxidized by metabolic enzymes such as alcohol dehydrogenase ADH and acetaldehyde dehydrogenase ALDH with the participation of coenzyme NAD+ to form NADH and acetic acid, accompanied by the production of byproducts of reactive oxygen free radicals (ROS); then, according to the needs of cells, acetic acid enters the tricarboxylic acid cycle or participates in protein post-translational modification. It is currently believed that acetaldehyde and ROS produced in the first stage are the main causes of fat accumulation and liver damage in ALD, and the second stage mainly disrupts post-translational modification and affects the long-term physiological function of the liver. Related research on the prevention and treatment of ALD focuses on reducing active molecules such as acetaldehyde and ROS produced by metabolism after alcohol intake, and reducing chemical damage to the liver caused by related molecules. For example, antioxidants (vitamin C, vitamin E, glutathione and its precursors, etc.), liver-protecting ingredients (silymarin, glycyrrhizic acid and bicyclol, etc.) and immunomodulators (interleukins, etc.) are used clinically. These drugs are used for people with liver function abnormalities with liver cell damage and elevated transaminases as the main manifestation, and are not recommended for people without elevated transaminases.

[0004] The development of new drugs to alleviate alcohol-induced liver damage is an urgent issue to be addressed. Summary of the invention

[0005] Based on this, the present application provides a composition comprising: Nicotinamide or its geometric isomers, tautomers, isotope-labeled substances, hydrates, solvates or pharmaceutically acceptable salts; and Tetramethylpyrazine (TMP) or its geometric isomers, tautomers, isotope-labeled substances, hydrates or solvates, and pharmaceutically acceptable salts.

[0006] In another aspect, the present application also provides a pharmaceutical preparation comprising the composition described herein.

[0007] In another aspect, the present application also provides use of the composition described herein or the pharmaceutical preparation described herein in the preparation of a medicament for treating and / or preventing a disease caused by nicotinamide adenine dinucleotide depletion.

[0008] In another aspect, the present application also provides use of the composition described herein in preparing food or health products.

[0009] In another aspect, the present application also provides a food or health product, which comprises the composition described herein.

[0010] In the past, the prevention and treatment of alcoholic liver damage mainly considered alleviating the chemical damage caused by acetaldehyde and ROS, while less attention was paid to the NAD+ consumption and metabolic disorders caused by alcohol. At present, substances such as NMN and NR cannot be used in medicines or health products, and NAM has problems with low bioavailability and potential toxic side effects. This application found that on the basis of a drug (tetramethylpyrazine) that promotes the remedial synthesis ability of NAD+, the combination of NAM can alleviate the degree of NAD+ depletion and metabolic disorders after drinking, and intends to fundamentally alleviate alcohol-induced liver damage. Experiments have found that the combination of TMP 50mg / kg, an ingredient that promotes NAD+ synthesis, and NAM 250mg / kg can effectively alleviate liver damage such as alcoholic lipidosis.

[0011] The composition of the present application can be used to prevent or alleviate liver damage such as liver metabolic disorders and liver steatosis caused by NAD+ consumption after drinking. The composition can be used as part of medical treatment, or it can be used for personal preference or lifestyle reasons. The present composition can be used as a dietary supplement, functional food, and medicine, etc. The present application provides a safe and effective method for improving the depletion of NAD+ in the liver after drinking and the various liver metabolic disorders caused by it, and has a good effect on the prevention and alleviation of alcoholic liver damage. The present application focuses on related molecules that promote the salvage synthesis of NAD+. In the case where only NAD+ synthesis precursors are used without using such molecules, there is the possibility of low bioavailability of the precursor molecules and potential side effects.

[0012] Other features and advantages of the present application will be described in the following description, and partly become apparent from the description, or be understood by implementing the present application. Other advantages of the present application can be realized and obtained by the schemes described in the description and the drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] The accompanying drawings are used to provide an understanding of the technical solution of the present application and constitute a part of the specification. Together with the embodiments of the present application, they are used to explain the technical solution of the present application and do not constitute a limitation on the technical solution of the present application.

[0014] Figure 1It is shown that TMP alleviates alcohol-induced liver damage under the NIAAA model in Example 1 of the present application. Among them, (A) H&E and Oil Red detect the effect of TMP administration on alcoholic liver pathological damage and lipid accumulation. (B) The effect of TMP administration on the biochemical indicators of alcoholic liver pathology, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and serum triglyceride (TG) content. Ctrl: control group; EtOH: alcohol group; EtOH+TMP 50mg / kg: 50mg / kg TMP is given while drinking; EtOH+TMP100mg / kg: 100mg / kg TMP is given while drinking. Compared with the ctrl group, *P<0.05, **P<0.01; compared with the EtOH group, #P<0.05, ##P<0.01.

[0015] Compared with the ctrl group, *P<0.05, **P<0.01; compared with the EtOH group, #P<0.05, ##P<0.01.

[0016] Figure 2 It is shown that in Example 2 of the present application, TMP alleviates the decrease in NAD+ content caused by alcohol, and the NAD+ / NADH ratio decreases. Among them, (A) TMP alleviates the decrease in NAD+ content caused by alcohol, NADH increases, and the NAD+ / NADH ratio decreases. (B) TMP cannot reduce NADH content (normalized F420 nM / F485 nM). (C) TMP alleviates the decrease in nicotinamide phosphoribosyltransferase (NAMPT) transcription level caused by alcohol. (D) TMP alleviates the decrease in NAMPT protein level caused by alcohol. Ctrl: control group; EtOH: alcohol group; EtOH+TMP 50mg / kg: 50mg / kg TMP is given while alcohol is given; EtOH+TMP 100mg / kg: 100mg / kg TMP is given while alcohol is given. Compared with the ctrl group, *P<0.05, **P<0.01; compared with the EtOH group, #P<0.05, ##P<0.01.

[0017] Figure 3It is shown that NAM combined with TMP in Example 3 of the present application alleviates alcohol-induced liver damage under the NIAAA model. Among them, (A) H&E and Oil Red detect the effects of NAM combined with TMP administration on alcoholic liver pathological damage and lipid accumulation. (B) The effects of NAM combined with TMP administration on biochemical indicators of alcoholic liver pathology, serum triglycerides, and total cholesterol (TCHO) content. Ctrl: control group; EtOH: alcohol group; TMP 50mg / kg: 50mg / kg TMP was given at the same time as alcohol; TMP+NAM (L / M / H): 50mg / kg TMP and low (L, 100mg / kg), medium (M, 250mg / kg), and high doses of NAM (H, 500mg / kg) were given at the same time as alcohol. Compared with the EtOH group, *P<0.05, **P<0.01; compared with the TMP group, #P<0.05, ##P<0.01. DETAILED DESCRIPTION

[0018] Unless otherwise stated, the technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which the application belongs. When a certain amount, concentration or other value or parameter is expressed in the form of a range, a preferred range, or a preferred upper numerical limit and a preferred lower numerical limit, it should be understood that it is equivalent to specifically revealing any range by combining any pair of upper range limits or preferred numerical values ​​with any lower range limit or preferred numerical value, regardless of whether the range is specifically disclosed. Unless otherwise stated, the numerical ranges listed herein are intended to include the endpoints of the range and all integers and fractions (decimals) within the range.

[0019] The terms "about" and "approximately" when used with a numerical variable generally refer to the value of that variable and all values ​​of that variable are within experimental error (e.g., within a 95% confidence interval of the mean) or within ±10% of the specified value, or a wider range.

[0020] The expression "comprising" or its synonymous similar expressions "including", "containing" and "having" etc. are open-ended and do not exclude additional unrecited elements, steps or ingredients. The expression "consisting of excludes any element, step or ingredient not specified. The expression "consisting essentially of means that the scope is limited to the specified elements, steps or ingredients, plus the optional elements, steps or ingredients that do not materially affect the basic and novel characteristics of the claimed subject matter. It should be understood that the expression "comprising" encompasses the expressions "consisting essentially of" and "consisting of".

[0021] The expression "at least one" or "one or more" means 1, 2, 3, 4, 5, 6, 7, 8, 9 or more.

[0022] Alcoholic liver injury is more likely to manifest as metabolic disorders without substantial liver damage, which is closely related to NAD+ consumption. The alcohol metabolism process consumes a large amount of intracellular NAD+ molecules to generate NADH. NAD+ is an important electron acceptor in cells and is widely involved in redox reactions and metabolic processes in cells. In the body, NADH is mainly restored to NAD+ through mitochondrial oxidative metabolism, but in the case of long-term drinking, the rate at which this mechanism consumes NADH is far less than the rate at which alcohol metabolism generates NADH, resulting in intracellular NAD+ depletion. NADH accumulation and NAD+ depletion can promote the development of ALD in many ways. NAD+ depletion can cause the weakening of ethanol metabolic enzyme activity, resulting in a large accumulation of toxic intermediates such as acetaldehyde. At the same time, NADH can spontaneously oxidize to generate ROS, causing cell damage. Excessive NADH can also change the intracellular metabolic process, promote fatty acid synthesis, promote the synthesis of unsaturated fatty acids, and then promote liver fat accumulation. Unsaturated fatty acids are susceptible to attack by active molecules such as ROS, generating peroxidized lipids. These peroxidation products can cause protein and DNA damage in cells in a chain reaction, promoting the occurrence and progression of ALD.

[0023] Supplementing NAD+ helps prevent and treat ALD. Literature has found that exogenous supplementation of NAD+ precursors such as nicotinamide mononucleotide (NMN), nicotinamide riboside (NR) and nicotinamide (NAM) can help prevent and treat alcoholic fatty liver. The applicant's previous research found that the fermented Chinese medicine light soybeans and its components NAM and tetramethylpyrazine (2,3,5,6-Tetramethylpyrazine, TMP) have a mitigating effect on ALD. TMP promotes the key NAD+ synthesis pathway - the salvage synthesis pathway. In general, the potential mechanism of the fermented Chinese medicine light soybeans in treating ALD is to relieve ALD through a two-pronged approach by supplementing NAD+ precursors and regulating the NAD+ synthesis pathway. Based on this, this application sets up different ratios of NAM and TMP, and explores their effects on alleviating ALD to obtain the optimal ratio of the NAD+ precursor that alleviates ALD and the TMP combination that regulates the components of the NAD+ synthesis pathway, providing data support for the application and development of this drug pair derived from fermented Chinese medicine ingredients.

[0024] The present application constructs a composition based on the fermentation product ingredients of light black beans, which includes precursors for NAD+ salvage synthesis and molecules that promote NAD+ salvage synthesis ability. The precursors for NAD+ salvage synthesis are selected from nicotinamide mononucleotide (NMN), nicotinamide riboside (NR) and nicotinamide (NAM). Among them, nicotinamide (NAM) is the most preferred. The molecule that promotes NAD+ salvage synthesis is selected from the light black bean ingredient tetramethylpyrazine (2,3,5,6-Tetramethylpyrazine, TMP) (chemical formula: C8H 12 N2) etc. Among them, tetramethylpyrazine (TMP) is the most preferred. Pharmaceutical NAD+ remedial synthesis precursors can be obtained in the form of dietary supplements or nutritional enhancers. TMP can be obtained from bacteria by biological fermentation.

[0025] In one aspect, the present application provides a composition comprising: Nicotinamide or its geometric isomers, tautomers, isotope-labeled substances, hydrates, solvates or pharmaceutically acceptable salts; and Tetramethylpyrazine (TMP) or its geometric isomers, tautomers, isotope-labeled substances, hydrates, solvates or pharmaceutically acceptable salts.

[0026] In some embodiments, the molar ratio of nicotinamide to tetramethylpyrazine is about 6: 1-1: 1. In some embodiments, the molar ratio of nicotinamide to tetramethylpyrazine is about 5: 1, 4: 1, 3: 1, 2: 1, or 1: 1. In some embodiments, the molar ratio of nicotinamide to tetramethylpyrazine is about 5.6: 1.

[0027] In some embodiments, other types of nicotinamide adenine dinucleotide (NAD+) precursors or their geometric isomers, tautomers, isotope labels, hydrates, solvates or pharmaceutically acceptable salts may also be used. In some embodiments, nicotinamide in the compositions described herein may be replaced by nicotinamide mononucleotide (NMN) or nicotinamide riboside (NR). In some embodiments, the molar ratio of nicotinamide adenine dinucleotide (NAD+) precursor to tetramethylpyrazine is about 6:1-1:1. In some embodiments, the molar ratio of nicotinamide adenine dinucleotide (NAD+) precursor to tetramethylpyrazine is about 5:1, 4:1, 3:1, 2:1 or 1:1. In some embodiments, the molar ratio of nicotinamide adenine dinucleotide (NAD+) precursor to tetramethylpyrazine is about 5.6:1.

[0028] In some embodiments, related molecules with similar functions can be obtained in other traditional Chinese medicines through similar experimental schemes as in the present application, or derivative modifications can be performed based on existing molecules in the present application, etc., to obtain new or similar NAD+-promoting salvage synthesis molecules.

[0029] In some embodiments, the mass ratio of nicotinamide to tetramethylpyrazine is about 7:1-1:1.

[0030] In some embodiments, the mass ratio of nicotinamide to tetramethylpyrazine is about 7: 1, 6: 1, 5: 1, 4: 1, 3: 1, 2: 1 or 1: 1. In some embodiments, the mass ratio of nicotinamide (NAM) to tetramethylpyrazine (2,3,5,6-Tetramethylpyrazine) is 5:1.

[0031] In another aspect, the present application also provides a pharmaceutical preparation comprising the composition described herein.

[0032] In some embodiments, the pharmaceutical formulation further comprises a pharmaceutically acceptable excipient selected from one or more of a granulating agent, a binder, a lubricant, a disintegrant, a sweetener, a glidant, an anti-adherent, an antistatic agent, a surfactant, an antioxidant, a gum, a coating agent, a colorant, a flavoring agent, a plasticizer, a preservative, a suspending agent, an emulsifier, an antimicrobial agent, a plant cellulose material or a spheronizing agent, and any combination thereof.

[0033] In some embodiments, the pharmaceutical formulations described herein also include pharmaceutically acceptable excipients. "Pharmaceutically acceptable excipients" means generally safe, nontoxic and desirable excipients that can be used to prepare pharmaceutical compositions, and include excipients that are acceptable for veterinary use and human medicine use. Such excipients can be solid, liquid, semisolid, or gaseous in the case of aerosol compositions. Pharmaceutical grade organic or inorganic carriers and / or diluents suitable for oral and surface use can be used to make compositions containing therapeutically active compounds. Diluents known in the art include aqueous media, vegetable oils and animal oils and fats. Stabilizers, wetting agents and emulsifiers, salts for changing osmotic pressure or buffers and skin penetration enhancers for ensuring appropriate pH values ​​can be used as adjuvants. Non-limiting examples of excipients include granulating agents, binders, lubricants, disintegrants, sweeteners, glidants, anti-adherents, antistatic agents, surfactants, antioxidants, gums, coating agents, colorants, flavoring agents, plasticizers, preservatives, suspending agents, emulsifiers, antimicrobials, plant cellulose materials, and spheronizing agents, and any combination thereof.

[0034] In some embodiments, the dosage form of the pharmaceutical preparation is selected from one or more of capsules, tablets, pills, liquids, powders, granules, fine granules, film coatings, pills, lozenges, sublingual agents, peptizers, buccal preparations, pastes, syrups, suspensions, elixirs, emulsions, coatings, ointments, plasters, poultices, transdermal preparations, lotions, inhalants, aerosols, injections, or suppositories.

[0035] In some embodiments, the compositions described herein or the pharmaceutical formulations described herein can be suitably prepared as pharmaceutical compositions and unit dosage forms for use as solids (e.g., tablets or filled capsules) or liquids (e.g., solutions, suspensions, emulsions, elixirs, or capsules filled therewith) for oral use, in the form of ointments, suppositories, or enemas for rectal administration, in the form of sterile injectable solutions for parenteral use (e.g., intramuscular, subcutaneous, intravenous, epidural, intraarticular, and intrathecal); or in the form of ointments, lotions, creams, gels, patches, sublingual strips or films for parenteral (e.g., topical, buccal, sublingual, vaginal) administration.

[0036] In some embodiments, a composition described herein or a pharmaceutical formulation described herein is administered orally, intravenously, intradermally, transdermally, intrathecally, intraarterially, intraperitoneally, intranasally, intravaginally, intrarectally, intravesically, intratumorally, topically, intramuscularly, subcutaneously, mucosally, by inhalation, injection, infusion, or any combination of the foregoing.

[0037] In some embodiments, a composition described herein or a pharmaceutical formulation described herein can be administered intravenously, intradermally, transdermally, intrathecally, intraarterially, intraperitoneally, intranasally, intravaginally, intrarectally, intravesically (e.g., such as directly administered into the bladder by injection or by intravesical instillation), intratumorally, topically, intramuscularly, subcutaneously, mucosally, orally, topically, by inhalation (e.g., aerosol inhalation), injection, infusion, continuous infusion, via a catheter, via lavage with an emulsifiable concentrate, by direct local infusion with a liquid composition (e.g., liposomes), or by other methods as known to one of ordinary skill in the art, or any combination of the foregoing.

[0038] In another aspect, the present application also provides use of the composition described herein or the pharmaceutical preparation described herein in the preparation of a medicament for treating and / or preventing a disease caused by nicotinamide adenine dinucleotide depletion.

[0039] In some embodiments, the disease caused by nicotinamide adenine dinucleotide depletion is alcoholism, alcoholic liver injury, or alcoholic liver disease.

[0040] In some embodiments, the disease caused by nicotinamide adenine dinucleotide depletion is alcoholic liver disease.

[0041] In some embodiments, the disease caused by nicotinamide adenine dinucleotide depletion is alcoholic steatosis.

[0042] In some embodiments, in addition to NAD+ depletion and metabolic disorders caused by drinking, there are also NAD+ depletion and metabolic disorders in the liver due to other causes such as diabetes, hyperlipidemia, poor eating habits, etc. The present application also has a mitigating effect on similar diseases with NAD+ disorders.

[0043] In another aspect, the present application also provides a method for treating and / or preventing a disease caused by nicotinamide adenine dinucleotide depletion, comprising administering a therapeutically effective amount of the composition described herein or the pharmaceutical preparation described herein to a subject.

[0044] In some embodiments, the therapeutically effective amount refers to administration of a composition described herein or a pharmaceutical formulation described herein such that the amount of nicotinamide adenine dinucleotide precursor (eg, NAM) and tetramethylpyrazine administered reaches 25-500 mg / kg and 25-100 mg / kg, respectively. In some embodiments, a therapeutically effective amount refers to administration of a composition described herein or a pharmaceutical formulation described herein such that the amount of nicotinamide adenine dinucleotide precursor (e.g., NAM) administered reaches 30 mg / kg, 40 mg / kg, 50 mg / kg, 60 mg / kg, 70 mg / kg, 80 mg / kg, 70 mg / kg, 100 mg / kg, 150 mg / kg, 200 mg / kg, 250 mg / kg, 300 mg / kg, 350 mg / kg, 400 mg / kg, 450 mg / kg, or 500 mg / kg; and the amount of tetramethylpyrazine administered reaches 30 mg / kg, 40 mg / kg, 50 mg / kg, 60 mg / kg, 70 mg / kg, 80 mg / kg, 90 mg / kg, or 100 mg / kg.

[0045] In some embodiments, the subject in the present application is a human, rat, mouse, cat, dog, horse, sheep, cow, monkey. In some embodiments, illustrative examples of subjects include primates, particularly humans, companion animals such as cats and dogs and similar animals, working animals such as horses, donkeys and similar animals, livestock animals such as sheep, cattle, goats, pigs and similar animals, laboratory test animals such as rabbits, mice, rats, guinea pigs, hamsters and similar animals, and captive wild animals such as captive wild animals in zoos and safari parks, deer, dingoes and similar animals. In some embodiments, the subject is a human.

[0046] In another aspect, the present application also provides use of the composition described herein in preparing food or health products.

[0047] In another aspect, the present application also provides use of the composition described herein in preparing a food or health product for preventing alcoholic liver injury.

[0048] In another aspect, the present application also provides a food or health product, which comprises the composition described herein.

[0049] In some embodiments, the food or health product further comprises an edible supplement.

[0050] In some embodiments, the food is a health food; or the food includes dairy products, soy products, meat products, or fruit and vegetable products; or the food is a beverage or a snack.

[0051] The rate at which alcohol metabolism consumes NAD+ to produce NADH is much faster than the rate at which NADH is converted into NAD+. At the same time, drinking alcohol also inhibits the body's ability to remediate NAD+ synthesis, further exacerbating the depletion of NAD+ in cells. Therefore, antagonizing the depletion of NAD+ caused by alcohol depends on exogenous supplementation.

[0052] Due to the unstable chemical properties of NAD+, NAD+ is currently supplemented in the body mainly by ingesting its precursors. Related molecules include NMN, NR and NAM. Among them, NMN and NR have not been approved by the US FDA and China NMPA as health products and medicinal uses. NAM has been maturely used as a medicine, health product and dietary supplement, but there are problems such as low utilization and toxic side effects in clinical practice. For example, long-term drinking inhibits the NAD+ salvage synthesis pathway, resulting in its substrate, NAM ingested into the body, unable to be converted into NAD+ in a timely and effective manner. In addition, excessive NAM can inhibit the enzymatic activity of metabolic enzymes that depend on NAD+. Excessive NAM can also be degraded by NNMT methylase, further reducing the bioavailability of NAM while consuming methylation donors, leading to epigenetic and signal transduction disorders in cells.

[0053] This application enhances the liver's NAD+ remedial synthesis capacity through drugs while supplementing NAM, reduces the degree of NAD+ depletion in the liver after alcohol intake, and alleviates liver metabolic disorders. This application can prevent and treat liver damage caused by drinking. In addition, the drug pair NAM and related traditional Chinese medicines have been used for many years and have shown that the drug is safe and effective, and the related ingredients are easy to extract and have mature industrial sources.

[0054] In summary, we considered giving NAD precursor NAM while enhancing NAMPT, a key enzyme in the NAD+ salvage synthesis pathway, to improve NAM bioavailability, so as to effectively increase the NAD+ content in the body and improve ALD. Nicotinamide (NAM) and 2,3,5,6-Tetramethylpyrazine (2,3,5,6-Tetramethylpyrazine) work together to maintain the homeostasis of nicotinamide adenine dinucleotide NAD+ content in the liver.

[0055] The present application describes multiple embodiments, but the description is exemplary rather than restrictive, and it is obvious to those skilled in the art that there may be more embodiments and implementations within the scope of the embodiments described in the present application. Although many possible feature combinations are shown in the drawings and discussed in the specific embodiments, many other combinations of the disclosed features are also possible. Unless specifically limited, any feature or element of any embodiment may be used in combination with any other feature or element in any other embodiment, or may replace any other feature or element in any other embodiment.

[0056] The present application includes and contemplates combinations of features and elements known to those of ordinary skill in the art. The embodiments, features and elements disclosed in the present application may also be combined with any conventional features or elements to form a unique invention scheme. Any features or elements of any embodiment may also be combined with features or elements from other invention schemes to form another unique invention scheme. Therefore, it should be understood that any feature shown and / or discussed in the present application may be implemented individually or in any appropriate combination. Therefore, except for the limitations made according to the attached claims and their equivalents, the embodiments are not subject to other restrictions. In addition, various modifications and changes may be made within the scope of protection of the attached claims.

[0057] In addition, when describing representative embodiments, the specification may have presented the method and / or process as a specific sequence of steps. However, to the extent that the method or process does not rely on the specific order of the steps described herein, the method or process should not be limited to the steps of the specific order described. As will be understood by those of ordinary skill in the art, other sequences of steps are also possible. Therefore, the specific sequence of the steps set forth in the specification should not be interpreted as a limitation to the claims. In addition, the claims for the method and / or process should not be limited to the steps of performing them in the order written, and those skilled in the art can easily understand that these sequences can be changed and still remain within the spirit and scope of the embodiments of the present application.

[0058] The experimental methods in the following examples without specifying specific conditions are usually measured according to national standards. The experimental materials in the following examples without specifying the source are all commercially available raw materials. The equipment used in each step in the following examples is conventional equipment. If there is no corresponding national standard, it is carried out according to the general international standards, conventional conditions, or according to the conditions recommended by the manufacturer. Unless otherwise defined or specified, all professional and scientific terms used in this application have the same meaning as those familiar to those skilled in the art. In addition, any method and material similar or equal to the recorded content can be applied to the method of this application.

[0059] Example The relevant materials used in the examples are as follows: 1. Reagents and Materials TMP (Chengdu Pufeide Biotechnology Co., Ltd.); nicotinamide NAM (Sigma-Aldrich Corporation, USA); Lieber-DeCarli liquid feed (Bio-Serv Corporation, USA); 42° Fenjiu (Shanxi Xinghuacun Fenjiu Factory Co., Ltd.); NAD+ / NADH detection kit (WST-8 method) (Shanghai Biyuntian Biotechnology Co., Ltd.).

[0060] 2. Instruments Liquid diet feeding tube with cap (Bio-Serv, USA); frozen section machine (Thermo Fisher, USA); disposable anticoagulant tube (Jiangsu Kangjian Medical Supplies Co., Ltd.); microplate reader (Synergy 2 Multi-Mode Microplate Reader, Bio Tek).

[0061] Example 1. Effects of promoting NAD+ salvage synthesis molecule TMP on alcoholic liver injury in chronic alcohol-induced liver injury mouse (NIAAA) model Experimental animals and dosing regimen: C57BL / 6J wild-type mice, male, 8-9 weeks old, were purchased from Sibeifu (Beijing) Biotechnology Co., Ltd. The animals were randomly divided into groups, with no less than 6 mice in each group, and first adapted to the control feed for 5 days. After the animals adapted, they were given control feed and liquid alcohol feed with a final concentration of 5%, and the relevant molecular drugs were given according to the group. TMP was gavaged with 0.5% sodium carboxymethyl cellulose solution at a dose of 50 mg / kg and 100 mg / kg. After 10 days, a single alcohol gavage (5g / kg) or isocaloric dextrin was given, and the mice were killed and dissected 9h after gavage. Serum and liver were collected for relevant analysis and testing. Experimental group settings: ctrl group: given control liquid feed; EtOH group: fed with liquid alcohol feed; TMP group: fed with liquid alcohol feed, and 50 mg / kg and 100 mg / kg TMP were given respectively.

[0062] Experimental protocol at the cell level: HEK293 cells were cultured in medium without pyruvate. The HEK293T NADH-SoNar detection system was constructed according to (Zhao, Yuzheng et al. Nature protocols. 11,8 (2016):1345-59.). HEK293 cells were transfected with SoNar expression plasmid and control plasmid and cultured for 48 hours. Subsequently, the transfected cells were seeded in 96-well plates. After 12 hours, the cells were given basal medium without pyruvate, medium containing 5 mM lactate (lactate, positive control), 5 mM pyruvate (pyruvate, negative control), and lactate + 100 mM TMP, respectively, and then cultured for 1, 2, 3, and 4 hours. The cells were rinsed twice, and the fluorescence intensity was detected by a microplate reader at 420 nm, 485 nm, and 528 nm excitation wavelengths.

[0063] Experimental results: The experimental results are shown in Figure 1 Compared with the ctrl group, the liver of the EtOH group mice had a large amount of lipid droplet accumulation, significant vacuolar changes, and inflammatory cell infiltration. The hepatocyte and tissue morphology of the mice in the TMP administration group were greatly improved compared with the model group. The number of lipid droplets in the low-dose TMP group decreased significantly and the volume decreased significantly. Among them, the morphology of the high-dose TMP group was relatively good. Except for the slight vacuoles, there was no significant change compared with the control group. The serum liver injury indicators AST and ALT of the mice in the administration groups were decreased in a dose-dependent manner compared with the model group (P<0.001, P<0.01, P<0.05). After the model mice were given TMP, low-dose (50 mg / kg) and high-dose (100 mg / kg) TMP could significantly reduce the serum TG of NIAAA mice (P<0.5, P<0.01).

[0064] Example 2. Effects of TMP-promoting NAD+ salvage synthesis molecules on alcohol-induced NAD+ levels and NAD+ synthesis in the NIAAA model Results Figure 2 , it can be seen that compared with the ctrl group mice, the NAD+ level of the EtOH group mice decreased, the NADH level increased slightly, and the liver NAD+ / NADH ratio decreased significantly (P<0.05). After TMP administration, the liver NAD+ level of mice in each group increased significantly, the NADH level decreased, and the liver NAD+ / NADH ratio increased significantly (P<0.05) ( Figure 2A in the figure). The decreasing trend of NADH is lower than the increasing trend of NAD+, suggesting that TMP regulates the NAD+ / NADH ratio by promoting NAD synthesis. To further verify the hypothesis, we detected the changes in the mRNA and protein levels of NAMPT, a key rate-limiting enzyme in NAD synthesis. The results showed that the changes in NAMPT protein levels were significantly decreased after alcohol administration ( Figure 2 C in the figure). The drug-treated group significantly reversed this trend. In addition, we used NADH-glo cell lines to test whether TMP could promote the conversion of NADH to NAD+. The results showed that TMP could not directly reduce NADH accumulation ( Figure 2 B). The above results suggest that TMP can act as a molecule to promote NAD+ salvage synthesis, increase NAD+ synthesis, and alleviate the damage of the NIAAA model.

[0065] Example 3. Effects of NAM combined with TMP on alcohol-induced liver injury in the NIAAA model Experimental animals and dosing regimen: C57BL / 6J wild-type mice, male, 8-9 weeks old, purchased from Sibeifu (Beijing) Biotechnology Co., Ltd. After random grouping, the animals were first adapted to the control feed for 5 days. After the animals were adapted, they were given control feed and liquid alcohol feed with a final concentration of 5%, and the relevant molecular drugs were given according to the group. NAM and TMP were dissolved in pure water and then gavaged. After 10 days of administration, a single alcohol gavage (5g / kg) or isocaloric dextrin was given. The mice were killed and dissected 9h after gavage. Serum and liver were collected for related analysis and testing. The NAM dosage was set as low (L), medium (M), and high (H) doses, and the L, M, and H doses were: 100 mg / kg, 250 mg / kg, and 500 mg / kg, respectively.

[0066] The ctrl group was fed with control feed; the EtOH group was fed with liquid alcohol feed; the TMP group was fed with liquid alcohol feed and 50 mg / kg TMP; the TMP+NAM group was fed with liquid alcohol feed and 50 mg / kg TMP and low (L), medium (M), and high (H) doses of NAM were given respectively.

[0067] Experimental results: The experimental results are as follows Figure 3As shown in the figure, compared with the model group, the hepatocyte edema of mice in the TMP monotherapy group was reduced, the fat droplets were significantly reduced, the cells were arranged more regularly and neatly, and the cytoplasm was filled. The model group was given TMP and NAM groups. As the dose of NAM increased, the hepatocyte edema and fat droplets were further reduced, the cells were arranged neatly, and the cytoplasm was filled. Among them, the subjective improvement in the TMP+NAM medium and high dose groups was more obvious. Compared with TMP alone, there was no significant decrease in serum ALT in mice in each TMP+NAM combined administration group, and AST and GGT were further decreased. In summary, the protective effect of TMP+NAM combined administration on alcohol-induced hepatocyte damage is stronger than that of TMP alone.

[0068] The lipid deposition in the liver of mice in each drug-treated group was significantly improved compared with the model group, among which the improvement in the medium and high dose groups of TMP+NAM was the most obvious, indicating that this dose of drug group has a good regulatory effect on the lipid metabolism of hepatocytes in NIAAA mice. On the basis of the single dose of TMP, TMP+NAM further reduced the serum TG and TCHO content of mice, among which the effect of the medium dose group of TMP+NAM group was the most significant (P<0.05), indicating that the drug has a positive regulatory effect on liver lipid metabolism. The combined administration of TMP+NAM has a synergistic effect.

[0069] In summary, the combination of TMP 50mg / kg, an ingredient that promotes NAD synthesis, and NAD precursor NAM 250mg / kg (the molar ratio of TMP to NAM is 1:5.6) can effectively alleviate liver damage such as alcoholic lipidosis.

[0070] Although the embodiments of the present application have been shown and described above, it can be understood that the above embodiments are exemplary and cannot be understood as limitations on the present application. Ordinary technicians in this field can change, modify, replace and modify the above embodiments within the scope of the present application.

Claims

1. A composition comprising: Nicotinamide or its geometric isomers, tautomers, isotope-labeled substances, hydrates, solvates or pharmaceutically acceptable salts; and Tetramethylpyrazine or its geometric isomers, tautomers, isotope-labeled substances, hydrates or solvates, and pharmaceutically acceptable salts.

2. The composition according to claim 1, wherein The molar ratio of the nicotinamide to the tetramethylpyrazine is about 6:1-1:

1.

3. The composition according to claim 2, wherein The molar ratio of the nicotinamide to the tetramethylpyrazine is about 5.6:

1.

4. A pharmaceutical preparation comprising the composition according to any one of claims 1 to 3.

5. The pharmaceutical preparation according to claim 4, wherein The pharmaceutical preparation further comprises a pharmaceutically acceptable excipient, which is selected from one or more of a granulating agent, a binder, a lubricant, a disintegrant, a sweetener, a glidant, an anti-adherent, an antistatic agent, a surfactant, an antioxidant, a gum, a coating agent, a colorant, a flavoring agent, a plasticizer, a preservative, a suspending agent, an emulsifier, an antimicrobial agent, a plant cellulose material or a spheronizing agent and any combination thereof.

6. The pharmaceutical preparation according to claim 4, wherein The dosage form of the pharmaceutical preparation is selected from one or more of capsules, tablets, pills, liquids, powders, granules, fine granules, film coatings, pills, lozenges, sublingual agents, peptizers, buccal preparations, pastes, syrups, suspensions, elixirs, emulsions, coatings, ointments, plasters, poultices, transdermal preparations, lotions, inhalants, aerosols, injections or suppositories.

7. Use of the composition according to any one of claims 1 to 3 or the pharmaceutical preparation according to any one of claims 4 to 6 in the preparation of a medicament for treating and / or preventing a disease caused by nicotinamide adenine dinucleotide depletion.

8. The use according to claim 7, wherein The disease caused by nicotinamide adenine dinucleotide consumption is alcoholism, alcoholic liver injury or alcoholic liver disease.

9. Use of the composition according to any one of claims 1 to 3 in preparing food or health products.

10. A food or health product comprising the composition according to any one of claims 1 to 3.

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