Field of the Invention

The present invention relates to compositions, in particular dietary compositions, that comprise a combination of choline cation and succinate anion (2-) and nicotinamide, and use of these compositions for maintaining and enhancing the brain energy metabolism in a human, and for preventing and treating symptoms and physiological conditions associated with an imbalanced, damaged or slowed brain energy metabolism in a human subject..

Background of the invention

Human body cells, especially in the brain, are dependent on oxygen for adenosine triphosphate (ATP) production in mitochondria. Nicotinamide adenine dinucleotide (NAD+) is a coenzyme that serves as a hydrogen carrier NADH/NAD+ between substrates of tricarboxylic acid cycle and complex I mitochondria supporting thus ATP production during the oxidative phosphorylation (Hinkle PC. Biochim Biophys Acta. 2005, 1706(1- 2):1-11. At normoxia (i.e. normal oxygen supply), NADH oxidation at mitochondrial complex I contributes to about 90% of total oxygen consumption (Lukyanova LD, Kirova Yl. Front Neurosci. 2015 9:320).

Nicotinamide (CAS No. 98-92-0) is a precursor of NAD+/IMADH and the vitamin B3. Nicotinamide is found in food and used as a dietary supplement. Nicotinamide is included in the list of essential medicines needed for health. (WHO Model List of Essential Medicines. World Health Organization, 2015). It has been demonstrated that nicotinamide incubation (5 mM) of fresh brain tissue slices 2 hr prior to hypoxia significantly increases total NAD(H) content, improves neuronal recovery, enhances ATP content, and prevents NADH hyperoxidation (Pavan K Shetty PK, Galeffi F., Turner DA. Neurobiol Dis. 2014, 62:469-478). Succinic acid or salts thereof (succinates) are substrates of mitochondrial complex II. At hypoxia, a condition of a deficiency in oxygen supply of tissues of the body, succinate oxidation may contribute to 70-80% of total oxygen consumption (Lukyanova LD, Kirova Yl. Front Neurosci. 2015 9:320). Because of high acidity (pKal=4.2 and pKa2=5.6), fumaric acid is dissociated completely at physiological range of pH 7.0-7.8 and is present in the body as dicarboxylate-anion (succinate). Succinate transport across the body is mediated by specific sodium-coupled dicarboxylate-anion transporters of SLC13 family. However, there is no specific transporting system for succinic acid or salts thereof across the blood-brain barrier (BBB), since no dicarboxylate-anion transporters were found in cerebral blood vessels or the choroid plexus (Chen X et al., J Clin Invest 1999, 103(8):1159-68). In line with this, it has been strictly proven that dicarboxylates cannot penetrate BBB in vivo (Hassel B et al., J Neurochemistry 2002, 82:410-419). Because of lack of transport capacity across the BBB, orally administered succinic acid and salts thereof generally are not effective in the treatment of disorders of central nervous system (CNS).

Choline salt of succinic acid (2:1) (hereinafter DISU or choline succinate (2:1) salt) is a specific salt of succinic acid that has been demonstrated to be surprisingly effective in the treatment of CNS disorders and enhancing cognitive function in animals (US 7,666,90; US 8,673,977). Further, it has been demonstrated that DISU salt is able to slow whole-brain ATP decline, in particular, it has a protective effect on the brain energy metabolism during global ischemia (Pomytkin I, Semenova N. Dokl Biochem Biophys 2005, 403:289-92). Treatment with DISU could preserve cognitive function in animals in a model of chronic cerebral hypoperfusion (Storozheva Z et al., BMC Pharmacol 2008, 8:1).

Choline is an essential nutrient for healthy metabolic functioning. Choline is needed for biosynthesis of acetylcholine, neurotransmitter crucial for communication of neurons in the nervous system. Choline deficiency leads to neurological disorders (Zeisel SH et a I, Nutr Rev. 2009, 67(ll):615-623), and oral administration of choline enhances cognitive function in relatively impaired performers (Knott V et al., Pharmacology, Biochemistry and Behavior 2015, 131:119-129). An adequate intake level for choline is 550 mg/day for men and 425 mg/day for women. The de novo choline synthesis is not sufficient to meet human requirements and choline must be obtained through the diet containing choline- rich foods, like eggs and shrimps, or as nutritional addition to a normal diet, for example in form of choline salts. Aging results in a physiological cerebral hypoxia, i.e. a deficiency in oxygen supply of the brain, due to a decrease of cerebral blood flow by approximately 28-50% from age of 30 to 70 years (Ogoh S and Ainslie PN. J AppI Physiol 2009, 107:1370-80). NAD ⁺ availability and abnormal NAD ⁺ /NADH redox state are also tightly linked to aging and age-related metabolic diseases and neurodegenerative disorders (Zhu X-H., et al Proc Natl Ac Sci USA 2014, 112:2876-2881). Based on results of several studies it has been suggested that complex II plays an important role in aging (see for review Wojtovich AP., et al. Biochem Biophys Acta 2013, 1827(5): doi:10.1016/j.bbabio.2012.12.007). In view of the fact that almost every country of the world at present is experiencing a shift in the distribution of a country's population towards older ages and huge socioeconomic problems resulting from the effects of an increasing environmental and psychological pressure on the general population, it is, therefore a great need in agents, that are safe and effective as both anti-aging nutritionals and pro-energetic dietary supplements for enhancing mental endurance and brain health in general population.

Summary of the invention

In a first aspect, the invention relates to a composition comprising, preferably, essentially consisting of choline cation, succinate anion (2-) and nicotinamide, or a nicotinamide derivate.

In a second aspect, the invention relates to a composition comprising, preferably essentially consisting of choline cation, succinate anion (2-) and nicotinamide, or a nicotinamide derivate for use in maintaining or enhancing the brain energy metabolism in a human.

In a third aspect, the invention relates to a composition comprising, preferably essentially consisting of choline cation, succinate anion (2-) and nicotinamide, or a nicotinamide derivate for use enhancing cognition, learning and/or memory, mental strength and endurance in a human.

In a forth aspect, the invention relates to a composition comprising, preferably essentially consisting of choline cation, succinate anion (2-) and nicotinamide, or a nicotinamide derivate for use in the dietary management of one or more symptoms and conditions associated with an imbalanced, damaged or reduced brain energy metabolism. In a fifth aspect, the invention relates to a method for the dietary management of a symptom or condition associated with an imbalanced, damaged or reduced brain energy metabolism in a human, or the dietary prevention of developing, occurring and/or re- occurring a symptom or condition associated with an imbalanced, damaged or reduced brain energy metabolism in a human, comprising administering to said human, at least once a day, a composition comprising, preferably essentially consisting of choline cation, succinate anion (2-) and nicotinamide, or a nicotinamide derivate.

The molar ratio of choline cation, succinate anion (2-) and nicotinamide, or the nicotinamide derivate, in the composition is preferably in the range from about 2:1:0.01 to about 2:1:10. The choline cation and succinate anion (2-) may derive from one and the same salt, in particular, choline succinate (2:1) salt, or from two different salts, e.g. choline bitartrate and succinate disodium salt. In one preferred embodiment, the composition is a nutritional composition or a part of a nutritional composition. Preferably, the composition is administered to a human daily in one or more doses for a period of one or more days.

The human may be an aging human individual and/or a human individual who is having or susceptible to psychological stress, fatigue, insomnia or mental depression.

One or more symptoms associated with an imbalanced, damaged or reduced brain energy metabolism in a human may be selected from one or more symptoms and conditions are selected from lack of mental concentration, weakened mental strength and endurance, frequent mood swings, mental depression, seasonal aftective disorder (SAD), insomnia, fatigue, weakened cognitive capacity, learning and memory.

In a sixth aspect, the invention related to use of a combination of choline cation, succinate anion (2-) and nicotinamide, or a nicotine derivate in a human, for restoring, maintaining and/or enhancing brain energy metabolism,

restoring, maintaining and/or enhancing mental and physical endurance;

maintaining and/or enhancing cognitive functions;

treating or reducing the risk of development or re-occurrence of a condition or a symptom associated with an imbalanced, damaged or reduced brain energy metabolism;

treating or reducing the risk of or delaying the onset of cognitive impairment associated with associated with an imbalanced, damaged or reduced brain energy metabolism

in said human.

The human may be any human individual, advantageously, the human is an aging human individual and/or a human individual who is having or susceptible to psychological stress, fatigue, insomnia or mental depression.

Detailed Description of the Invention

All terms and definitions explained throughout the text of specification of the invention relate to all aspects and embodiments of the invention, unless otherwise specified. The present invention relates to synthetic compositions that are safe and effective to maintain and enhance energy metabolism, advantageously, the brain energy metabolism in humans. The compositions can be used to support or re-establish a proper brain energy metabolism and to help to enhance the physical and mental endurance in a human, including healthy human individuals and individuals diagnosed with a disease, without age limitation and in a combination with any diet and/or therapy. According to the invention, the compositions comprise at least three compounds that are associated with the energy metabolism, preferably, the compositions essentially consist of three compounds, choline cation and succinate anion (2-) and nicotinamide (NAM), preferably in a molar ratio from about 2:1:0.01 to about 2:1:10; advantageously, the choline cation and succinate anion are present in the composition in a form of a choline succinate salt, preferably choline succinate (2:1) salt (interchangeably termed herein "di-choline succinate" or "DISU". The term "(2:1)" in the present context means a single molecule of the choline succinate salt comprises two choline cations and one succinate anion (2-). In some embodiments, the invention relates to a derivate of NAM as an alternative NAM in the composition of the invention.

It was surprisingly found by the inventors that a synthetic composition essentially consisting of choline cation, succinate anion (2-) and nicotinamide, exogenously added to mammalian brain cells in vitro is synergistically effective for increasing the level of both nicotinamide adenine dinucleotide (NADH) and adenosine triphosphate (ATP) in the cells. An especially enhanced production of NADH and ATP in mitochondria of the brain cells was observed when the choline cation and succinate anion (2-) were derived from di-choline succinate salt (DISU) present in the composition. However, a significant enhancement of the production of ATP and NADH could also be achieved when the choline cation and succinate anion of the composition were derived from other salts of comprising said cation and anion. Furthermore, the inventors found that human individuals treated with an aqueous composition essentially comprising DISU and NAM observed at least one of the following effects: reduced symptoms of seasonal affective disorder (SAD), decreased symptoms of fatigue, frequency and scale of mood swings, phycoiogicai depression and stress, and, as contrary, an increased prevalence of good mood and well -being feeling, better mental concentration, clarity and endurance of mind. Moreover, there have been recorded a persistent increase in the level of cellular energy deport phosphocreatine (PCr) in the brain of a human individual in vivo following two weeks of intake of a beverage essentially consisting of the composition of the invention, i.e. a combination of DISU and NAM.

The wording "composition essentially consisting of" <certain compounds> means that the certain compounds of the composition are essential and the only compounds of the composition that are responsible for the biological effect associated with the composition. Thus, the wording does not exclude other compounds that do not directly contribute to the effect. The term "synthetic" in the present context means a man-made composition. The synthetic compositions of the invention may comprise both synthetically prepared molecules that are structurally identical to the molecules that naturally occurring in the living bodies, and artificial molecules that do not have natural structural equivalents. The term "about" means a deviation from the indicated value by 0,01% to 10%, such as as from 0,5% to 5%. The term "choline cation" means the cation having the chemical formula C ₅ H _i4 NO ^' (CAS No. 62-49-7). The term "succinate anion" means succinic acid anion (2-) having the chemical formula C H ₄ 0 (CAS No. 110-15-6}, The terms "di-choline succinate", "choline succinate salt (2:1)" and "DiSU" are interchangeable and mean the molecule of formula (I):

The term "nicotinamide" or " NAM" means the molecule identified with CAS No. 98-92- 0. The term "derivate of nicotine amide" of NAM derivate" means a molecule that is derived from NAM by a synthetic process, i.e. NAM is a start molecule for the synthesis of the derivate, e.g. nicotinamide riboside (CAS No: 1341-23-7) or nicotinamide mononucleotide (CAS No: 1094-61-7). As mentioned above, a first aspect of the invention relates to a composition comprising, preferably essentially consisting of choline cation, succinate anion (2-) and NAM, or a NAM derivate, preferably the choline cation and succinate anion are present in the molar ratio 2:1, preferably, said cation and anion are present in form of choline succinate salt (2:1) (DISU), i.e. DISU is a part of the composition. Alternatively, the choline cation of the composition may derive from another salt of choline, e.g. choline bitartrate (CAS No. 87-67-2), and succinate anion may derive from another salt of succinic acid, e.g. succinic acid disodium salt (CAS No. 6106-21-4). Compositions essentially consisting of choline bitartrate, succinic acid disodium salt and NAM may be preferred in some embodiments of the invention. As mentioned, the molar ratio of choline cation, succinate anion (2- and NAM in the composition of the invention may vary from about 2:1:0.01 to about 2:1:10. Compositions with the molar ratio of the individual compounds within this range acts synergetic and enhance the production of major energy molecules ATP, NADH and PCr in the brain cells. Further, these compositions are efficient in dietary management of symptoms of seasonal affective disorder (SAD), mood swings, insomnia, fatigue, weakened mental concentration and endurance, phycological stress and depression in human individuals, the symptoms are typically associated with imbalanced or weakened energy metabolism. Some preferred embodiments of the ratio of the compounds in the composition are described in the working examples, however, the exemplified compositions are not-limiting embodiments of the invention as the composition has the energy generating effect described herein through the whole ratio range.

To obtain the above effects, when used for the purposes described herein, the essential compounds of the composition, i.e. choline cation, succinic acid anion (2-) and NAM, or a NAM derivate, are present in so called "effective amounts". The effective amounts of the compounds may vary depending on the aim and/or method of use, and on the subject in need. These embodiments are discussed below and exemplified by non-limiting working examples.

In some embodiments, a composition of the invention comprising choline cation, succinate anion (2-) and NAM may further comprise creatine (CAS No.57-00-1), or a creatine precursor, such as e.g. amino acids glycine and arginine. Other useful additives to a composition comprising choline cation, succinate anion (2-) and NAM, or a NAM derivate, are discussed below. Compositions of the invention can be advantageously used for maintaining, establishing or re-establishing a proper level of brain energy metabolism in a human, wherein "proper level of brain energy metabolism" means a dynamic capability of the brain cells to generate energy by producing biological "energy" molecules NADH, ATP and PCr in the amounts that are sufficient to maintain normal brain functioning, including the processes relating to cognition, learning, memory, mental focus and endurance, etc, under normal conditions, and in the amounts that can compensate for an increased energy consumption by the brain processes under extraordinary conditions, e.g. in situations characterized by an extraordinary overload or complexity of mental tasks, or situations demanding an increased mental and/or physical endurance and/or performance, etc. The proper level of brain energy metabolism may differ from one human individual to another, and may depend on age, education level, social or health status, mental or physical capability of the individual. The proper level may be defined according to existing standards for mental performance, e.g. using Cognitive Tests and Performance Validity Tests https://www.ncbi.nlm. nih.gov/books/NBK305230/).

The expression "establishing/re-establishing/restoring the proper level of brain energy metabolism" means in the present context that the composition provides an extraordinary support in generation of energy, i.e. generation of energy-providing molecules NADH, ATP and PCr, to an energy-depleted and weakened brain (e.g. due to a disease, age, environmental or psychological factors). For the maintaining, establishing or re-establishing a proper level of brain energy metabolism in a human, the composition can be used as an everyday dietary supplement both by healthy individuals and medical patients, as it is safe to use in a combination with any diet and medicaments. Advantageously, in one embodiment, the composition may be used for maintaining or enhancing the brain energy metabolism in an aging human. The term "aging human" in the present context generally relates to a human individual of 14 years old or older, preferably, a human individual over 24-25 years old. In another embodiment, the composition can be advantageously used for the prophylaxis of ageing of the brain. It is well known that the energy generating capability of the brain gradually diminishes with ageing. Used on everyday basis for maintaining a proper brain energy metabolism by a human individual at earlier age, including infants and young children, the composition of the invention has an advantageous capability to maintain the speed and complexity of energy-demanding processes of the brain unchanged through the life span of the individual, or, at least, it can decrease the speed of brain ageing associated with the depletion of energy deports by persistently fueling the ageing brain with energy and allowing the ageing brain to perform on the level of a young brain. The compositions of the invention may be formulated as nutritional compositions. Preferably, said compositions comprise effective amounts of the essential ingredients of the composition of the invention in as nutraceuticals, preferably in an appropriate molar ratio as described above. The term "nutraceutical" means a pharmaceutical-grade and standardized nutrient. The term "nutrient" means in the present context substance that provides nourishment essential for the maintenance of life of a human. The term "nutritional" in the present context means that the composition is for the dietary supplementation of a human individual. The term "dietary supplement" means a product taken by mouth that contains a dietary ingredient, e.g. a nutrient, intended to supplement the diet. The term "human" in the context of the invention relates to any human individual. In one preferred embodiment, the human is an aging human, such as a human individual over 2Λ-.25 years old, for example over 30 years old, such as between 35 and 55 years old, or between 40 and 60 years old, etc. In another preferred embodiment, the human is an individual who is having or susceptible to a psychological stress, fatigue, insomnia or mental depression. In one preferred embodiment, the human is an aging individual who is having or susceptible to a psychological stress, fatigue, insomnia or mental depression. The amounts of choline cation, succinate anion (2-) and NAM,, e.g. DISU and NAM, in a composition of the invention may be adjusted for use by a particular individual or a group of individuals according to the individual's needs, age, physiological conditions, etc., and depending on the dosage form and administration regime For example, the amount of NAM in the composition may vary from 10 mg to 4000 mg per serving, served as one or more dosages a day, such as about 25-2000 mg per serving, served as several dosages per day, or about 50-1000 mg per serving served as several dosages per day, etc, wherein the daily dose of NAM will depend on the dietary demand of a concrete human individual or a group of human individuals. Some dietary compositions useful for some specific demands are described in non-limiting working examples of the invention. However, a daily intake of about 4000 mg of NAM in the composition is considered as safe and effective for any described herein purpose. The amount of DISU per serving may vary from 10 mg to 1000 mg per serving, and it can be served as one or more dosages a day. Accordingly, to archive a desired effect, as any of the described herein, without having any negative physiological effects, an individual may intake a composition comprising up to 4000 mg NAM, or a NAM derivate, and up to 1000 mg DISU, or the corresponding amounts of choline cation and succinate (2-) anion derived from other salts of choline and succinic acid, daily. In one preferred embodiment, a nutritional composition of the invention comprises essentially DISU and NAM, and the molar ratio of choline cation, succinate anion (2-) and nicotinamide in the composition is about 2:1:0.4, correspondingly. In another preferred embodiment, the molar ratio of choline cation, succinate anion (2-) and nicotinamide in the composition is about 2:1:1. The term "about" in the present context means a 1-10% deviation from the indicated value. Preferably, the intake is continuous for a period of at least one week or, preferably, for a longer period of time, such as 2 to 4 weeks, 1 to 12 months or longer. There is practically no limitation for how long the composition can be intaken as a dietary supplement. The intake can be interrupted any time for a period of time and resumed again, when the individual feels that it is needed, e.g. in connection with changes in individual's life, health, seasonal fluctuations of individual's physiological/mental state, or age. A dietary manager of ordinary skill can readily determine the amounts of said ingredients of the dietary composition according to the accepted rules and regulations. In further aspects, the invention relates to a composition as any of the described above (i) for use in restoring, maintaining or enhancing the brain energy metabolism in a human; (ii) for use in enhancing concentration, attentiveness, mental strength and endurance, cognition, learning and/or memory in a human; (iii) for use in treating and/or reducing the symptoms of phycological stress, lack of mental concentration, weakened mental strength and endurance, frequent mood swings, mental depression, seasonal affective disorder (SAD), insomnia, fatigue; (iv) for use in delaying the onset of brain ageing associated with a decline of the brain capability of generating energy molecules, such as NADH, ATP and PCr, necessary for energy-demanding processes relating to performing complex metal tasks, cognition, learning, memory, attentiveness, focus, etc. The term "mental strength" means the ability of a human individual to regulate the individual's thoughts, control emotions, and behave productively despite the circumstances. The term "mental endurance" means the ability to exercise mental strength in everyday life, and the ability to deal effectively with all challenges. Accordingly, use of a combination of choline cation, succinate anion (2-) and nicotinamide, or a nicotine derivate, as described herein, in a human, for

restoring, maintaining and/or enhancing brain energy metabolism,

restoring, maintaining and/or enhancing mental and physical endurance; maintaining and/or enhancing cognitive functions;

treating or reducing the risk of development or re-occurrence of a condition or a symptom associated with an imbalanced, damaged or reduced brain energy metabolism;

- treating or reducing the risk of or delaying the onset of cognitive impairment associated with associated with an imbalanced, damaged or reduced brain energy metabolism

in said human is beneficial and safe, independently whether this combination a part of a composition comprising a broad spectrum other compounds (e.g. such as described below) or as a solo composition essentially comprising a salt of choline, providing the choline cation, a salt of succinic acid, providing the succinate (2-) anion, and nicotine amine or a nicotine amide derivate as described herein.

The term "cognitive impairment" in the present context means "mild cognitive impairment (MCI)" that is characterized by a slight but noticeable and measurable decline in cognitive abilities, including memory and thinking skills. A person with MCI is at an increased risk of developing Alzheimer's or another dementia. The term "restoring" is used interchangeably with the term "re-establishing" and means bringing back to or putting back into a former or original state The invention also includes aspects relating to methods of dietary management of symptoms and conditions associated with imbalanced, damaged or reduced brain energy metabolism, lack of mental concentration, weakened mental strength and endurance, frequent mood swings, mental depression, seasonal affective disorder (SAD), insomnia, fatigue, weakened cognitive capacity, learning and memory, said methods comprising a step of administration of a composition according to the invention to a human individual in need. The term "dietary management" in the present context means the practice of providing a nutritional option for individuals and groups with health concerns instead of a therapeutic intervention or as a prophylactic treatment, preferably under supervision of a dietary or medical professional. Advantageously, the compositions of the invention can be used for in dietary prophylaxis of primary and/or secondary development or occurrence of a symptom or condition associated with an imbalanced, damaged or reduced brain energy metabolism in a human, including reducing the risk of occurrence and re-occurrence of the same symptom or condition, and reducing the strength and duration of the symptom or condition that is occurred for the first time or is re- occurring. The term "primary" means that the symptom or condition occurs in the human for the first time; the term "secondary" means that the symptom or condition is reoccurring. Accordingly, a method for the dietary prevention of development, occurrence and/or re-occurrence of a symptom or condition associated with an imbalanced, damaged or reduced brain energy metabolism in a human, wherein said symptom or condition may be any of the described herein, is one of the aspects of the invention The term wording "imbalanced, damaged or reduced" in the present context means that an individual's brain energy metabolism is not on a proper level (as discussed above), but is weakened by a disease, physiological or phycological condition of the individual, harmful environmental factors. As mentioned above, in some embodiments the invention relates to nutritional compositions comprising nutrients other than the essential compounds of the composition. A nutritional composition of the invention may be in form of a nutritional product including without limitations a food, a beverage, a dietary supplement, a functional food, and a medical food. In one preferred embodiment, a composition of the invention is an aqueous nutritional composition, e.g. a drink or beverage.

In practicing the invention, the compounds of the composition of the invention can be prepared by any process known in the art or obtained from a known commercial manufacturer, e.g. nicotinamide or its derivatives, choline bitartrate, succinate disodium salt, may be obtained from Merck. DISU can be prepared by the reaction of choline hydroxide (CAS No. 123-41-1) with succinic acid (CAS No.110-15-6) as, e.g. described in WO2009/022933A1.

The nutritional compositions described herein may be prepared by procedures well- known from the art and may contain some other optional ingredients. Such optional ingredients generally are used individually at levels from about 0.0005% to about 10.0% by weight of the composition. Examples of suitable optional ingredients include, but are not limited to, carriers, minerals, carbohydrates, lipids, vitamins, co-factors, buffers, flavors and sweeteners, inorganic salts, cations and anions typically abandoned in natural drinking water, taste modifying and/or masking agents, carbon dioxide, amino acids, organic acids, antioxidants, preservatives, and colorants. The nutritional compositions can be combined with one or more carriers and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, chewing gums, foods, beverages, and the like.

Non-exclusive examples of ingredients which can serve as carriers include water; sugars, such as glucose, lactose, and sucrose; cellulose, and its derivatives; starches, such as corn starch and potato starch; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter; oils, such as olive oil, peanut oil, cottonseed oil, corn oil and soybean oil; glycols, such as propylene glycol; esters, such as ethyl oleate and ethyl laurate; polyols, such as glycerin, mannitol, sorbitol, and polyethylene glycol; agar; buffering agents; water; pH buffered solutions; and other non-toxic compatible substances employed in formulations. Wetting agents, emulsifiers and lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, release agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the compositions. Non-exclusive examples of antioxidants are Vitamin E, ascorbic acid, carotenoids, aminoindoles, Vitamin A, uric acid, flavonoids, polyphenols, herbal antioxidants, melatonin, lipoic acids, and mixtures thereof.

Non-exclusive examples of useful inorganic salts typically abandoned in natural drinking water are sodium carbonate, sodium bicarbonate, potassium chloride, magnesium chloride, calcium chloride, and mixtures thereof.

Non-exclusive examples of useful cations are sodium, potassium, magnesium, calcium, zinc, iron, and mixtures thereof. Non-exclusive examples of useful anions are fluoride, chloride, bromide, iodide, carbonate, bicarbonate, sulfate, phosphate, and mixtures thereof. Non-exclusive examples of suitable buffers are phosphate buffer, glycine buffer, citrate buffer, acetate buffer, carbonate buffer, tris-buffer, triethanolamine buffer, and succinate buffer. Non-exclusive examples of suitable flavors are synthetic flavor oils; flavoring aromatics and naturals oils such as cinnamon oil, oil of wintergreen, peppermint oils, clove oil, bay oil, anise oil, eucalyptus, thyme oil, cedar leave oil, oil of nutmeg, oil of sage, oils of citrus fruits, oil of bitter almonds, and cassia oil; plant extracts, flowers, leaves, fruits, vanilla, chocolate, mocha, coffee, apple, pear, peach, citrus such as lemon, orange, grape, lime, and grapefruit; mango, strawberry, raspberry, cherry, plum, pineapple, and apricot, and combinations thereof.

Non-exclusive examples of suitable sweeteners are natural and synthetic sweeteners. Non-exclusive examples of natural sweeteners are naturally occurring substances, sucrose, extracts from naturally occurring substances; extracts of the plant Stevia ebaudiana Compositae Bertoni such as stevia, steviol, rebaudiosides A-F, and dulcosides A and B; extracts of Thladiantha grosvenorii such as mogroside V and related glycosides and triterpene glycosides; phyllodulcin and its derivatives; thaumatin and its derivatives; mogrosides such as mogroside IV, mogroside V, siamenoside, and mixtures thereof; genus Siraitia including S. grosvenorii, S. siamensis, S. silomaradjae, S. sikkimensis, S. Africana, S. borneesis, and S. taiwaniana; naturally-occurring glycosides; and active compounds of plant origin having sweetening properties, and mixtures thereof. Non-exclusive examples of synthetic sweeteners are aspartame saccharin, and mixtures thereof.

Non-exclusive examples of suitable colorants are dyes suitable for food such as those known as FD&C dyes, natural coloring agents such as grape skin extract, beet red powder, titanium dioxide, and beta-carotene, annatto, carmine, chlorophyll, paprika, and mixtures thereof.

Non-exclusive examples of useful organic acids are acetic acid, butyric acid, malic acid, pyruvic acid, glutamic acid, citric acid, omega-3 unsaturated acids, linoleic acid, linolenic acid, eicosapentaenoic acid, docosahexaenoic acid, aspartic acid, and mixtures thereof. Non-exclusive examples of useful amino acids are Glycine, Arginine, L-Tryptophan, L- Lysine, Methionine, Threonine, Levocarnitine, and L-carnitine. Non-exclusive examples of useful vitamins are thiamin, riboflavin, nicotinic acid, panthothenic acid, biotin, folic acid, pyridoxine, vitamin B12, lipoic acid, vitamin A, vitamin D, vitamin E, ascorbic acid, choline, carnitine; alpha, beta, and gamma carotenes; vitamin K, and mixtures thereof. Non-exclusive examples of useful co-factors are thiamine pyrophosphates, flavin mononucleotide, flavin adenine dinucleotide, pyridoxal phosphate, biotin, tetrahydrofolic acid, Coenzyme A, coenzyme B12, 11-cis-retinal, 1,25- dihydroxycholecalciferol and mixtures thereof. In one embodiment a nutritional composition of the invention may comprise compounds that are able to increase the blood circulation, e.g. an extract of Ginkgo biloba or ginseng. In some embodiments, a composition of the invention may comprise an antioxidant, e.g. astaxanthin, resveratrol, flavonoids. As mentioned, the nutritional compositions can be used as a component of a food product.

Nonexclusive examples of food products include regular foods, dietary supplements, beverages, and medical foods.

The term "medical food" refers to a food which is formulated to be consumed or administered enterally under the supervision of a physician and which is intended for the specific dietary management of a disease, condition, or disorder. Preferably, the nutritional compositions are administered to a human orally for a period of at least one day or, preferably, longer (as discussed above). The following non-limiting working examples and are presented to illustrative the invention. Accordingly, embodiments described in the working examples are not limiting the scope of the invention in any way. Example 1. Embodiments of nutritional compositions of the invention

Beverage 1. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water to provide a beverage.

Beverage 1

This beverage (molar ratio choline:succinate:NAM is 2:1:0.4), when administered orally to a subject daily, 330 ml per serving, one or more servings, is useful for improvement of brain energy metabolism in a subject, helping the subject to maintain high mental endurance and persistent feeling of well-being.

Beverage 2. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water.

This beverage (molar ratio choline:succinate:NAM is 2:1:2), when is administered orally to a subject daily, 330 ml per serving, one or more servings, is useful for dietary management of disorders accompanying cerebral hypoxia and psychological stress. Beverage 3. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water.

Beverage 3 Ingredient Content, per serving

NAM 188 mg

DISU 500 mg

Succinic acid qs to pH 6.5

Water to 330 ml

This beverage (molar ratio choline:succinate:NAM is 2:1:1), when is administered orally to a subject daily, 330 ml per serving, one or more servings, is useful for improvement of brain energy reserve, as measured by phosphocreatine-to-ATP ratio. Beverage 4. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 500 ml of water.

Beverage 4

Ingredient Content, per serving

NAM 210 mg

DISU 560 mg

Water to 500 ml

This beverage (molar ratio choline:succinate:NAM is 2:1:1), when is administered orally to a subject daily, 500 ml per serving, one or more servings, is useful for balancing the brain energy to the level that is necessary for healthy brain functioning.

Beverage 5. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water.

Beverage 5

Ingredient Content, per serving

NAM 375 mg

DISU 100 mg

Succinic acid qs to pH 6.5

Water to 330 ml

This beverage (molar ratio choline:succinate:NAM is about 2:1:10), when is

administered orally to a subject daily, 330 ml per serving,one or more servings, is useful for enhancing of brain healthy functioning. Beverage 6. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water.

Beverage 6

Ingredient Content, per serving

NAM 7 mg

DISU 2000 mg

Succinic acid qs to pH 6.5

Water to 330 ml

This beverage (molar ratio choline:succinate:NAM is about 2:1:0,01), when administered orally to a subject daily, 330 ml per serving, one or more servings, is useful for enhancing of brain healthy functioning.

Beverage 7. The beverage is prepared by mixing of NAM with DISU in amounts as indicated below and dissolved in 330 ml of water to provide a beverage.

Beverage 7

Ingredient Content, per serving

NAM 37 mg

DISU 250 mg

Citric acid qs to pH 6.5

Water to 330 ml

This beverage (molar ratio choline:succinate:NAM is 2:1:0.4), is to be administered orally to a subject daily, 330 ml per serving, one or more servings, is useful for improvement of brain energy metabolism in a subject, helping the subject to maintain high mental endurance and persistent feeling of well-being.

Example 2. Evaluation of the effect of DISU on energy metabolism in ischemia damaged brain in vivo.

Male rats received 10 mg/kg DISU (n=8), 10 mg/kg disodium succinate (n=8), 10 mg/kg dipotassium succinate (n=8), 10 mg/kg choline chloride (n=8), or saline (control, n=23) for 7 days. Then, global ischemia was induced in rats by cardiac arrest under anesthesia and whole-brain ATP levels were measured by ³¹ P NM in vivo for 15 min. Results are presented in Table 3 as meaniSEM ATP levels in the brain at 15 ^th min after ischemia onset, in percent to baseline (accepted for 100%). Table 1

*Differs significantly from saline (p<0.05, f-test).

The data of Table 1 show that DISU protects the brain from ATP decline at global hypoxia as compared to the control, while DISU constituents, choline and succinate, taken individually and in the same doses, were not effective. Notably, DISU protection of brain energy metabolism was significant, while another succinate salt, disodium succinate, was not effective. Also, another choline salt, choline chloride, was not effective in protection of brain energy metabolism. Thus, DISU significantly differs from other choline and succinate salts in its capacity to protect brain energy metabolism in ischemic conditions. Notably, DISU did not affect ATP levels in the brain under normoxia (Pomytkin I, Semenova N. Dokl Biochem Biophys 2005, 403:289-92).

Example 3. Evaluation of the effect of a composition of the invention on the production of ATP in brain cells in vitro.

Materials and methods. Mixed primary cultures of cortical neurons and glial cells were prepared from mouse pups 1-3 days postpartum, as described by Vaarmann A. et al., 2010 (Cell Calcium 2010, 48(2-3):176-82). For measurements of ATP, the cells at 14-21 DIV were transfected for 24 h with the ATP sensing probe ATI.03 (as described in Imamura H et al., PNAS 2009, 106(37):15651-15656) using Lipofectamine 2000 according to the manufacturer's instructions. In experiments, cells were exposed to 50 μΜ DISU, 20 μΜ NAM, nicotinic acid (niacin) (20 μΜ), composition of 50 μΜ DISU and 20 μΜ NAM, composition of 50 μΜ DISU and 20 μΜ nicotinic acid, or none (control) for about 5 min. Then, ATP levels in cells were measured using confocal microscopy. In another series of experiments, cells were exposed to reference compounds including sodium succinate (50 μΜ), choline bitartrate (100 μΜ), or compositions thereof and the ATP levels in cells have also been obtained for comparison. All data presented were obtained from at least 5 coverslips and 2-3 different cell preparations.

Results. Data are presented in Table 2 as mean of ATP levels in cells. Mean of ATP levels in non-treated cells (control) was accepted for 100%. Effects of the treatments on ATP levels in cells (ΔΑΤΡ) were calculated by formula: ΔΑΤΡ = ATP _treat - ATP _contr0 |, wherein ATP _t r _eat is ATP level in treated cells and ATP _contr0 | is ATP levels in control cells.

Table 2

Treatment ATP, % ΔΑΤΡ, %

Control 100 0

DISU 242* +142

Sodium succinate 123 +23

NAM 114 +14

Nicotinic acid 101 +1

Choline bitartrate 140 +40

DISU and NAM" 328* +228

DISU and Nicotinic acid" 252* ^† +52

Sodium succinate and NAM" 142 +42

Sodium succinate and NAM and Choline bitartrate" 246 * ^† +146

*Differs significantly from control (p<0.05).

"Differs significantly from DISU (p<0.05).

†Differs significantly from composition"DISU and NAM" (p<0.05).

Table 2 shows that

treatment with the composition consisting of DISU and NAM results in a significant synergistic increase in intracellular ATP levels by 228% compared to the control (p<0.05). The effect of the composition ΔΑΤΡ = 228% was 1.5-fold greater than sum of the effects ΔΑΤΡ = 142 + 14 = 156% of the individual components, DISU and NAM, respectively. There is a significant difference between the effect ΔΑΤΡ = 228% of the composition and the effect ΔΑΤΡ = 142% of DISU alone (p<0.05). Thus, the composition of the invention is synergistically effective for the improvement of ATP production in brain cells; treatment with the composition of DISU and Nicotinic acid results in an increase in intracellular ATP levels by 152.0% compared to the control (p<0.05). There is no significant difference between the effect ΔΑΤΡ = 152% of the composition and the effect ΔΑΤΡ = 142% of DISU alone (p>0.05). The sum of the individual effects ΔΑΤΡ = 142 + 1 = 143% of DISU and Nicotininc acid is similar by value to the effect ΔΑΤΡ = 152% of the composition. Thus, the combined effect of the compounds of the composition on the production of ATP is non-synergistic. Despite of NAM and nicotinic acid (niacin) are generally considered as members of vitamin B3 family with similar biological capabilities, the presented data clearly show that their effects on the production of ATP in brain cells when they are combined with DISU are different. treatment with the composition of Sodium succinate and NAM and Choline bitartrate resultes in a significant synergistic increase in intracellular ATP levels by 146% compared to the control (p<0.05). The effect of the composition ΔΑΤΡ = 146% was about 1.8-fold greater than sum of the effects ΔΑΤΡ = 23 + 14 + 40 = 77% of its individual components, Sodium succinate, NAM, and Choline bitartrate. Thus, the composition of Sodium succinate, NAM, and Choline bitartrate was also synergistically effective for the improvement of ATP production in brain cells. The data indicate, that the presence of choline anion in a composition comprising a succinate salt and NAM is important for the synergetic generation of ATP in mitochondria: there is a significant difference between the effect of the composition "Sodium succinate and NAM and Choline bitartrate" (ΔΑΤΡ = 146% ) and the effect of the composition "Sodium succinate and NAM" (ΔΑΤΡ = 42%) (p<0.05). Thus, the composition of the invention essentially comprising succinate anion, choline cation and NAM is synergistically effective for the production of ATP in brain cells; a choice a combination of succinate anion and choline cation is important for the scale of the synergetic effect of the composition of the invention: there is a significant difference between compositions "DISU and NAM" and "Sodium succinate and NAM and Choline bitartrate" in their effects on ATP production in the brai n cells (p<0.05): the composition "DISU and NAM" was about 1.6-fold more effective than "Sodium succinate and NAM and Choline bitartrate" despite on the same molar concentrations of choline cation, succinate anion, and NAM in the compositions. It indicates that DISU is a superior representative for the combination of choline anion and succinate cation. - treatment individually with Sodium succinate and NAM, or a composition

thereof, results in a non-significant increase in intracellular ATP levels compared to the control (p>0.05), by 23%, 31%, or 42%, respectively. The sum of the individual effects ΔΑΤΡ = 23 + 31 = 54% of Sodium succinate and NAM is similar by value and slightly greater, compared to the effect of the composition , ΔΑΤΡ = 42%. Therefore, the composition of sodium succinate and NAM is not synergistically effective for the ATP production in the brain cells, which is in sharp contrast to the composition of the invention, comprising succinate anion, NAM and choline cation. Accordingly, we conclude that a composition of invention essentially comprising choline cation, succinate anion and NAM at the molar ratio about 2:1:04 is effective for the enhancement of ATP production in the brain cells.

Example 4: Evaluation of the effect of a composition of the invention of the production of NADH in brain cells in vitro

NADH is a donor of electrons for electron transport chain in mitochondria. The level of NADH in mitochondria is a balance between production of this molecule in TCA cycle and consumption of it in complex I. Measurements of NADH in live cells (tissue) can be done using autofluorescence of NADH (excitation 360nm, emission ~430nm). However, the mitochondrial signal of NADH could not be separated from NADPH autofluorescence or from the cytosolic NADH signal (Bartalome F and Abramov AY Methods Mol Biol. 2015, 1264:263-70).

Methods and Results

Mixed cultures of cortical neurons and glial cells were prepared as described (Vaarmann et al., 2010), from mouse pups 1-3 days postpartum. Neurons were easily distinguishable from glia: they appeared phase bright, had smooth rounded somata and distinct processes, and positioned just above the focal plane of the glial layer. Cells were used at 14-21 DIV. Experiments were conducted in HBSS solution at the 37° C. To estimate the mitochondrial pool of NADH, maximal activation of mitochondrial respiration is induced by uncoupler FCCP (mitochondrial NADH is taken 0) followed by inhibition of respiration by NaCN (NADH is maximal due to inhibition of consumption). This set up provides evaluation of several characteristics: the rate of NADH production in TCA cycle (recovery of the NADH signal after NaCN), total mitochondrial pool and redox index.

Application of sodium succinate (50μΜ) to primary neurons and astrocytes induced fast and significant decrease of the mitochondrial NADH level which corresponds to activation of complex I -related respiration (n=156 cells, N=3 experiments). This is a typical effect of the addition of succinate which needs to be compensated by the following NADH production (Reeve et a I., Cell Death Dis. 2015 Jul 16;6:el820). DISU in contrast to sodium succinate induced only slight initial decrease in mitochondrial NADH followed by a slow steady increase in NADH autofluorescence (to 123±6 % of control after 20 min of incubation). There were not detected a noticeable increase in NADH autofluorescence after 20 min of incubation with sodium succinate.

Adding a combination of NAM (20μΜ) and DISU (50μΜ) (the molar ratio choline:succinate:NAM is about 2:1:0.4) not only reduced initial drop in NADH, but significantly increased i NADH autofluorescence detected in 20 min following the initial drop (161±11% of control; n=132; N=3 experiments). Application of NAM (20μΜ) alone in short term (15-20 min) induced only moderate increase (to 116±8% of control) in mitochondrial NADH pool (N=3; n=68 cells).

Accordingly, a combination of DISU and NAM has a significantly higher effect of the generation of NADH in brain mitochondria than the single compounds of the composition, and that the effect of the individual compounds is synergetic

Based on the above results, we conclude that a composition essentially comprising DISU and NAM is effective for the generation of NADH in the brain cells.

Example 5. Evaluation of the effect of intake of a beverage comprising DISU and NAM on the level of phosphocreatine in human brain in vivo ATP is strongly buffered in the brain by conversion via creatine kinase-catalysed reaction to phosphocreatine (pCr), the phosphorylated analogue of the guanidino amino acid creatine. ATP can be resynthesized from phosphocreatine 12 times faster than via oxidative phosphorylation and more than 70 times faster than de novo pathways. Phosphocreatine (pCr) is a high energy substrate that serve as a reserve ("temporal energy buffer") in the brain to recycle adenosine triphosphate (ATP), the direct energy source for energy-consuming reactions in the cell (Wallimann T et al., Biochem J 1992, 281 (Pt l):21-40; Valenzuela et al Brain Res Rev. 2007,56:198-213; Rae et al, Proc. R. Soc. Lond. B. 2003, 270:2147-2150).

Methods

A volunteer (57 years old healthy female subject,) received a beverage containing 500 mg of DISU and 188 mg NAM (the molar ratio choline:succinate:NAM is about 2:1:1) in 100 ml of water, once-a-day for seven consecutive days. The volunteer has been maintaining the normal diet and did not receive any other food supplements during the trail. Whole-brain levels of PCr and ATP were obtained using the ³¹ P magnetic resonance spectroscopy ( ³¹ P MRS). ³¹ P MRS at 3 Tesla (3T) at day 0 (just before the beverage intake), day 7 and day 16 (following the beverage intake). The ratio pCr-to-γΑΤΡ signals (pCr/ATP) in ³¹ P MRS spectra was used as a measure of the pCr levels in the brain. At every session, ³¹ P M RS spectra were recorded in three replicates (n=3); signals related to pCr, ATP were evaluated and the pCr-to-ATP ratio were calculated. Effect of the beverage intake on pCr/ATP ratios (ApCr/ATP) in the brain was calculated by formula: ApCr/ATP = 100% (mean pCr/ATP _day7/16 - mean pCr/ATP _day0 )xl00%/mean pCr/ATP _day0 . indicates significant between-groups mean differences in f-Student test (p<0.05).

Results

The results of the ³¹ P MRS measurements are presented in Table 3. The data are presented as mean ± sem (n=3) for pCr, ATP, pCr/ATP, and ApCr/ATP.

Table 3

*Differs significantly from a value at day 0 (p<0.05). As it can be seen from the data presented in Table 3, intake of the beverage containing DISU and NAM, where the molar ratio choline:succinate:NAM is about 2:1:1, results in a significant increase in the level of brain phosphocreatine (PCr) after 7 days of the consumption that continues to increase by the 16 ^th day of consumption, by 7 % and 13% compared to baseline value (at day 0), respectively. No significant changes in brain ATP levels to the baseline (day 0) were recorded neither at day 7 (p=0.48) or day 16 (p=0.57), indicating that a steady-state ATP levels in the brain are stable.

Accordingly, we conclude that the composition of the invention is effective in supporting the brain energy metabolism and energy reserves necessary for complex mental activity in any human individual, and, particular, in an ageing individual which mental activity is slowed and decreased.

Example 6. Evaluation of the intake of a beverage comprising DISU and NAM: human trail

Study design

A total of 20 male and female healthy adults (21-63 years old) were recruited to participate in the study. The participants were randomized into 2 groups, each of 10 subjects. First group was receiving beverage A (see Table 4 below). Second group was receiving beverage B (placebo product). The placebo and treatment products were bottled beverages in the 500 ml dosage container.

Table 4

Participants were distributed and filled the study questionnaire (see below). They were also received a supply of the study beverages in doses, one dose per day for continued consumption for the period of 12 weeks, and copies of the study questionnaire. Participants were instructed to consume the products during the day before 18:00 and reminded to avoid consuming more than 250 mg caffeine and less than 2,5 g taurine per day during the study period. The participants were instructed to fill the questionnaire (1) in the end of the twelfth week following the start of the consumption of the beverage. In the end of the study (end of the twelfth week) the participants delivered the questionnaire and, additionally, they were interviewed regarding their general experience of the beverage (distributed and filled the beverage evaluation questionnaire (2) (see below)). Results of the study.

The symptom evaluation (questionnaire 1).

The study was run during 12 weeks through November, December and January.

1 Often = daily or several times a day and without an obviously serious reason

1 Seldom = sometimes and due to a reason, reason being not obvious

'' Never = time to time and only due to an obvious reason

The beverage experience evaluation (questionnaire 2)

From the data of questionnaire 1 it can be concluded that physical condition of subjects of Group A did in general improve as they experienced a change from "worse to better" in all of the assessed symptoms. In the same time, there were not observed much changes in the symptoms during the trail in the subjects of group B. As most of the symptoms, if not all, are generally associated with lack of energy in the brain or unbalance in the brain energy metabolism, it can be concluded that the composition of the invention is useful to maintain a healthy level of or to enhance the brain energy metabolism in general public that is daily exposed to a vast amount of socio- environmental stimuli which are demanding mental energy to deal with, leading to exhaustion of the brain energy reserves and, consequently, to development of the above symptoms .