Compositions and methods using a combination of oleuropein and fisetin for cellular energy

Abstract

A composition is provided, comprising a combination of oleuropein or its metabolites and fisetin or its derivatives.The composition can be an oral nutritional composition, such as a dietary supplement, an oral nutritional supplement, a food product, or a food for special medical purposes (FSMP).The composition can be administered to an individual in need thereof for (i) improving physiological conditions associated with metabolic fatigue in one or more cells, (ii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells, and (iii) increasing antioxidant capacity, reducing oxidative stress and / or enhancing mitochondrial function, and (iv) treating or preventing calcium deficiency / depletion disorders in an individual.In addition or alternatively, the method can treat or prevent mitochondrial-related diseases or conditions associated with altered mitochondrial function in an individual in need of treatment or at risk of such treatment.

Description

[Background technology]

[0001]

[0001] The present disclosure relates generally to compositions and methods for managing energy at the cellular level using a combination of oleuropein or its metabolites and fisetin or its metabolites. In some embodiments, the compositions and methods can enhance mitochondrial function in middle-aged or elderly individuals and increase bioenergetics through activation of the mitochondrial calcium uniporter, thus promoting cellular activation.

[0002]

[0002] Population ageing is a significant demographic phenomenon. As the growth of the elderly population exceeds the growth of the total population due to longer life spans, the proportion of the elderly population relative to the rest of the population has increased significantly, partly due to declining birth rates. For example, in the 1950s, one in every 12 people was aged 60 or over; by the end of the 2000s, one in every 10 people was aged 60 or over. It is projected that by the end of the 2050s, one in every five people worldwide will be aged 60 or over.

[0003]

[0003] Age-related changes in brain morphology and cerebrovascular function are commonly observed in middle-aged and elderly people, who often suffer from some degree of cognitive impairment, including progressive decline in physical function and / or cognitive function. Cognitive decline has been consistently reported in conjunction with aging in a wide range of cognitive domains, including processing speed, attention, episodic memory, spatial ability, and executive function. Brain imaging studies have found that these normal age-related cognitive declines are associated with a decrease in both gray and white matter volume in the brain, with the fronto-striatal system being the most severely damaged with age. Such cortical volume loss may be due to many deleterious cellular processes associated with normal aging, such as the long-term accumulation of damage from free radicals resulting in oxidative damage, chronic low-grade inflammation, accumulation of homocysteine ​​(which, if increased, is a risk factor for cognitive impairment and dementia), and reduced mitochondrial function. In addition to direct cellular damage, the brain also suffers indirect damage from damage to the microvasculature. It is clear that the pathology of aging, and even dementia, involves a complex interplay between these interrelated factors. For example, mitochondrial dysfunction results in increased oxidative stress, which can trigger inflammation and vascular damage.

[0004]

[0004] Mitochondria are the primary source of aerobic energy production in mammalian cells and also maintain a large Ca2+ gradient across the inner membrane, providing the signaling potential for the molecule. In addition, mitochondrial Ca2+ may play a role in regulating ATP production in mitochondria and contribute to orchestrating cellular metabolic homeostasis. (Glancy, B. et al. (2012). "Role of mitochondrial Ca2+ in the regulation of cellular energetics." Biochemistry 51(14):2959-2973). Alterations in mitochondrial Ca2+ homeostasis have been associated with various pathological conditions and are important in the pathogenesis of several human diseases (Arduino et al. Journal Physiol. 2018 Jul;596(14):2717-2733).

[0005]

[0005] In recent years, nutrition, education, exercise and cognitive exercise have been demonstrated as potential interventions to prevent age-related decline in physical and cognitive function. A wealth of clinical, epidemiological and individual evidence supports the role of individual nutritional factors in reducing dementia risk and age-related neurodegeneration. However, formal trials of nutritional interventions have yielded mixed results (Schmitt et al., Nutrition Reviews 68:S2-S5 (2010)). In addition, although there have been efforts to harness the mitochondrial Ca2+ transport mechanism for therapeutic intervention, there are currently no pharmacological compounds that induce and selectively regulate mitochondrial Ca2+ homeostasis. Summary of the Invention

[0006]

[0006] In view of the experimental data disclosed later in this specification, the present inventors believe that the combination of oleuropein and fisetin improves the efficiency of mitochondrial energy production.

[0007]

[0007] Thus, in a general embodiment, the present disclosure provides a composition comprising a therapeutically effective amount of oleuropein and / or metabolites in combination with fisetin and / or derivatives for use in improving a physiological condition associated with metabolic fatigue in one or more cells, (ii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells, and (iii) increasing antioxidant capacity, reducing oxidative stress and / or enhancing mitochondrial function, and (iv) treating or preventing calcium deficiency / depletion disorders in an individual.

[0008]

[0008] In another embodiment, the present disclosure provides a composition comprising a therapeutically effective amount of oleuropein and / or its metabolites in combination with fisetin and / or derivatives for delaying the onset of metabolic decline, maintaining muscle mass and / or muscle function, reducing oxidative stress, maintaining immune function, and / or maintaining cognitive function in healthy middle-aged and older adults.

[0009] In a further embodiment, the present disclosure also comprises: i) enhancing at least one of mental or muscular performance in an individual; or ii) providing a composition comprising a combination of oleuropein and / or its derivatives and fisetin and / or its derivatives in a therapeutically effective amount for improving or maintaining cognitive function in an individual.

[0010]

[0010] In another embodiment, the present invention provides a unit dosage form comprising a combination of oleuropein and / or derivatives thereof and fisetin and / or derivatives in effective amounts for at least one of: (i) treating, reducing the incidence, or reducing the severity of a mitochondrial-related disease or condition associated with altered mitochondrial function; (ii) improving a physiological condition associated with metabolic fatigue in one or more cells; (iii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells; and (iv) treating or preventing calcium deficiency / depletion disorders; (v) increasing metabolic rate; (vi) improving or maintaining cognitive function; and (vii) enhancing or maintaining mitochondrial function.

[0011]

[0011] In another embodiment, the invention provides a kit comprising, in one or more containers, a combination of oleuropein and / or its metabolites in combination with fisetin and / or derivatives.

[0012]

[0012] Advantages and additional features will become apparent from the following drawings and detailed description. [Brief description of the drawings]

[0013] [Figure 1] 1 shows the chemical structures of fisetin (A) and oleuropein (B). [Diagram 2]1 is a graph showing that the effect of the combination of fisetin and oleuropein is greater than that of fisetin or oleuropein alone on mitochondrial activation by increasing mitochondrial Ca2+ in HeLa cells. The bar graph shows the effect of oleuropein (3 μM, black), fisetin (3 μM, gray), and the combination of 3 μM fisetin + 3 μM oleuropein on the integrated mitochondrial calcium rise induced by 100 μM histamine. Results are expressed as mean ± SEM of n=6 experiments. * indicates statistically significant difference at P<0.05 (one-way ANOVA test) for the difference between measured and theoretical values ​​of mitochondrial calcium. [Diagram 3] 1 is a graph showing that fisetin synergizes with oleuropein to activate mitochondria by increasing mitochondrial Ca2+ in HeLa cells. To quantify the synergistic effect of the combination of oleuropein and fisetin on mitochondrial activation, the expected theoretical effect (sum of fisetin effect and oleuropein effect, extracted from data in FIG. 2) was compared with the actual measured effect of the combination (fisetin+oleuropein, extracted from data in FIG. 2). Results are expressed as mean±SEM of n=6 experiments. * indicates statistical significance at P<0.05 (Student's t-test) for the difference between measured and theoretical values ​​in mitochondrial calcium. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014]

[0016] definition

[0017] Some definitions are provided below. However, definitions may be found in the "Embodiments" section below, and the heading "Definitions" above does not imply that such disclosure in the "Embodiments" section is not a definition.

[0015]

[0018] All percentages stated herein are by total weight of the composition unless otherwise indicated. As used herein, "about," "approximately," and "substantially" are understood to refer to numbers within a certain range of numerical values, for example, within the range of -10% to +10% of the referenced number, preferably within the range of -5% to +5% of the referenced number, more preferably within the range of -1% to +1% of the referenced number, and most preferably within the range of -0.1% to +0.1% of the referenced number. All numerical ranges herein should be understood to include all integers or fractions within the range. Furthermore, these numerical ranges should be interpreted as supporting claims directed to any number or subset of numbers within the range. For example, a disclosure of 1 to 10 should be interpreted as corresponding to ranges of 1 to 8, 3 to 7, 1 to 9, 3.6 to 4.6, 3.5 to 9.9, etc.

[0016]

[0019] As used in this disclosure and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context dictates otherwise. Thus, for example, reference to "a metabolite" or "the metabolite" includes one metabolite, but also includes two or more metabolites.

[0017]

[0020] The terms "comprise", "comprises", and "comprising" should be interpreted as inclusive rather than exclusive. Similarly, the terms "include", "including", and "or" should all be interpreted as inclusive unless such interpretation is clearly prevented by the context. However, the compositions disclosed herein may not include elements not specifically disclosed herein. Thus, disclosure of an embodiment using the term "comprising" includes disclosure of an embodiment "consisting essentially of" the specified components, and an embodiment "consisting of" the specified components.

[0018]

[0021] As used herein, "a composition essentially comprising at least one oleuropein or metabolite thereof" and "a composition essentially comprising calcium and at least one oleuropein or metabolite thereof" do not include any additional compounds that affect mitochondrial calcium transport other than at least one oleuropein or metabolite thereof and optional calcium. In certain non-limiting embodiments, the composition consists of an additive, at least one oleuropein or metabolite thereof, and optionally calcium.

[0019]

[0022] The term "and / or" used in the context of "X and / or Y" should be interpreted as "X" or "Y" or "X and Y". Similarly, "at least one of X or Y" should be interpreted as "X" or "Y" or "both X and Y". For example, "at least one oleuropein or a metabolite thereof" means "oleuropein" or "a metabolite of oleuropein" or "both oleuropein and a metabolite thereof".

[0020]

[0023] As used herein, the terms "example" and "such as," especially when followed by a list of terms, are merely exemplary and illustrative and should not be considered as exclusive or inclusive. As used herein, "associated with" and "linked with" mean occurring simultaneously, preferably caused by the same underlying condition, and most preferably, one of the specified conditions is caused by the other specified condition.

[0021]

[0024] The terms "food," "food product," and "food composition" refer to a product or composition intended for ingestion by an individual, such as a human, and providing at least one nutrient to such an individual. The compositions of the present disclosure, including the many embodiments described herein, may comprise, consist of, or essentially comprise the elements disclosed herein, as well as any additional or optional ingredients, components, or elements described or not described herein that are useful in the diet.

[0022]

[0025] As used herein, the terms "treatment" and "treating" refer to the administration of a composition disclosed herein to a subject having a condition to attenuate, reduce or ameliorate at least one symptom associated with the condition and / or to slow, reduce or prevent the progression of the condition. The terms "treatment" and "treating" include both preventative or prophylactic treatments (treatments that prevent and / or slow the onset or progression of the targeted pathological condition or disorder) and curative, therapeutic or disease-modifying treatments, including, for example, therapeutic measures to cure, delay, reduce symptoms and / or halt the progression of a diagnosed pathological condition or disorder, as well as treatment of patients at risk of or suspected of having the condition, and patients who are unwell or diagnosed as having a disease or medical condition. The terms "treatment" and "treating" do not necessarily mean treating the subject until they are cured. The terms "treatment" and "treating" also refer to maintaining and / or enhancing the health of an individual who is not afflicted with a disease but who may be susceptible to an unhealthy condition. The terms "treatment" and "treating" are also intended to include the synergism or otherwise enhancement of one or more primary preventative or therapeutic measures. As non-limiting examples, treatment can be performed by the patient, a caregiver, a doctor, a nurse, or another medical professional.

[0023]

[0026] Both human and animal treatments are within the scope of this disclosure. Preferably, at least one oleuropein or metabolite thereof is administered in a serving or unit dosage form that provides a therapeutically or prophylactically effective amount.

[0024]

[0027] The terms "prevent" and "prevention" refer to administering a composition disclosed herein to a subject not exhibiting any symptoms of the condition to reduce or prevent the onset of at least one symptom associated with the condition. Additionally, "prevention" includes reducing the risk, incidence, and / or severity of a condition or disorder.

[0025]

[0028] As used herein, an "effective amount" is an amount that treats or prevents a deficiency, treats or prevents a disease or medical condition in an individual, or, more generally, reduces symptoms, manages the progression of a disease, or provides a nutritional, physiological, or medical benefit to an individual.

[0026]

[0029] The relative terms "improved," "increased / enhanced / enhanced," "enhanced / potentiated," and the like refer to the effect of the compositions disclosed herein, i.e., compositions comprising an effective amount of at least one oleuropein or metabolite thereof, compared to administration of an otherwise identical composition lacking oleuropein and lacking oleuropein metabolites over the same period of time.

[0027]

[0030] As used herein, "administering" includes providing the referenced composition to an individual by another individual so that the individual can ingest the composition, and also simply includes the individual's own act of ingesting the referenced composition.

[0028]

[0031] "Animal" includes mammals, including but not limited to rodents, aquatic mammals, domestic animals such as dogs, cats, and other pets, livestock such as sheep, pigs, cows, and horses, and humans. When "animal", "mammal", or plurals thereof are used, these terms also apply to any animal in which, depending on the context, a demonstrated or intended effect can be demonstrated, for example, by the animal benefiting from improved mitochondrial calcium transport. Although the term "individual" or "subject" is often used herein to refer to a human, the disclosure is not so limited. Thus, the term "individual" or "subject" refers to any animal, mammal, or human that can benefit from the methods and compositions disclosed herein.

[0029]

[0032] The term "pet" refers to any animal that can benefit from or enjoy the compositions provided by the present disclosure. For example, a pet may be an animal such as a bird, bovine, canine, equine, feline, caprine, wolf, murine, ovine, or porcine, although a pet may be any suitable animal. The term "companion animal" refers to a dog or cat.

[0030]

[0033] A "subject" or "individual" is a mammal, preferably a human. The term "elderly", in relation to humans, means an age of at least 60 years, preferably greater than 63 years, more preferably greater than 65 years, and most preferably greater than 70 years. The term "older adult" in the human context means a postnatal age of 45 years or more, preferably greater than 50 years, more preferably greater than 55 years, and includes elderly people. The term "older adult" in the human context means a postnatal age of 45 years or more, preferably greater than 50 years, and more preferably greater than 55 years, and includes elderly people.

[0031]

[0034] As used herein, "frailty" is defined as a clinically discernible state of heightened vulnerability resulting from age-related declines in reserve and function across multiple biological systems, resulting in impaired ability to cope with routine or acute stressors. The presence of one or two of these criteria identifies the pre-frailty stage as being at high risk for progression to frailty.

[0032]

[0035] As used herein, the terms "serving" or "unit dosage form" are interchangeable and refer to a physically separate unit suitable as a unitary dose for human and animal subjects, each unit containing a predetermined amount of a composition comprising at least one oleuropein or metabolite thereof as disclosed herein, preferably with a pharma- ceutically acceptable diluent, carrier, or vehicle, in an amount sufficient to produce a desired effect. The specifications of the unit dosage form depend on the specific compound used, the effect to be achieved, and the pharmacodynamics associated with each compound in the host. In one embodiment, the unit dosage form may be a predetermined amount of liquid contained in a container such as a bottle.

[0033]

[0036] An "oral nutritional supplement" or "ONS" is a composition that contains at least one macronutrient and / or at least one micronutrient, e.g., in a sterile liquid, semisolid, or powder form, and is intended to supplement other nutritional intakes, such as those from food. Non-limiting examples of commercially available ONS products include MERITENE®, BOOST®, NUTREN®, and SUSTAGEN®. In some embodiments, the ONS can be in a liquid form that can be consumed without the addition of additional liquid, e.g., the amount of liquid is one serving of the composition.

[0034]

[0037] As used herein, "incomplete nutrition" refers to a nutritional product that preferably does not contain sufficient levels of macronutrients (protein, fat and carbohydrate) or micronutrients to be the sole source of nutrition for the animal to which it is administered. The term "complete nutrition" refers to a product that can be the sole source of nutrition for a subject. An individual can obtain 100% of its nutritional requirements from a complete nutritional composition.

[0035]

[0038] "Kit" means that the components of the kit are physically associated in or with one or more containers and are considered a single unit for manufacture, distribution, sale, or use. Containers include, but are not limited to, bags, boxes, cartons, bottles, packaging of any type or design or material, overwraps, shrink wraps, affixed components (e.g., stapled or glued), or combinations thereof.

[0036]

[0039] "Metabolic fatigue" refers to a decrease in mitochondrial function in one or more cells (e.g., one or more of the liver, kidney, brain, and skeletal muscle) due to a lack of substrate in one or more cells and / or accumulation of metabolic products in muscle fibers, which inhibits either calcium release or the ability of calcium to stimulate mitochondrial function. Physiological conditions associated with metabolic fatigue may include muscle fatigue or weakness, lack of energy, particularly lack of physical energy, lack of vitality, or muscle weakness.

[0037]

[0040] Embodiment

[0041] The present disclosure provides compositions comprising a therapeutically effective amount of oleuropein and / or its metabolites in combination with fisetin and / or derivatives for use in (i) improving a physiological condition associated with metabolic fatigue in one or more cells, (ii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells, and (iii) increasing antioxidant capacity, reducing oxidative stress and / or enhancing mitochondrial function, (iv) treating or preventing calcium deficiency / depletion disorders in an individual.

[0038]

[0042] Oleuropein is a polyphenol found in the fruits, roots, stems and more specifically the leaves of plants belonging to the Oleaceae family, in particular the olive (Olea europaea).

[0039]

[0043] In one embodiment, at least a portion of the oleuropein is obtained by extraction, for example from one or more of the stems, leaves, fruits, or kernels of a plant belonging to the family Oleaceae, preferably from a plant belonging to the family Oleaceae, such as olive (olive tree), plants of the genus Ligustrum, plants of the genus Syringa, plants of the genus Fraximus, plants of the genus Jasminum, plants of the genus Osmanthus, etc. Additionally or alternatively, at least a portion of the oleuropein and / or metabolites can be obtained by chemical synthesis.

[0040]

[0044] Non-limiting examples of suitable metabolites of oleuropein include oleuropein aglycone, hydroxytyrosol, elenolic acid, homovanillyl alcohol, isohomovanillyl alcohol, glucuronidated forms thereof, sulfated forms thereof, derivatives thereof, and mixtures thereof.

[0041]

[0045] Fisetin (7,3',4'-flavone-3-ol) (see Figure 1) is a polyphenol found in many plants and functions as a yellow / ochre colorant. Fisetin is also found in many fruits and vegetables such as strawberries, apples, persimmons, grapes, onions, and cucumbers.

[0042]

[0046] In one embodiment, at least a portion of the fisetin is obtained by known means, such as by extraction from a plant / vegetable / fruit source of fisetin. Additionally or alternatively, at least a portion of the fisetin and / or metabolites may be obtained by chemical synthesis.

[0043]

[0047] Non-limiting examples of suitable metabolites of fisetin include its glucuronidated form, its sulfate form, its derivatives, and mixtures thereof. In a preferred embodiment, the derivative is gerardol.

[0044]

[0048] Fisetin may be derived from any suitable source and may be isolated and / or chemically synthesized.

[0045]

[0049] In a preferred embodiment, oleuropein and fisetin and derivatives are obtained from plant sources. For example, oleuropein can be obtained from olive plants. For example, fisetin can be obtained from strawberries, apples, persimmons, grapes, onions, cucumbers, etc.

[0046]

[0050] The composition may also include one or more additional bioactive compounds, such as one or more compounds selected from the group consisting of antioxidants, anti-inflammatory compounds, glycosaminoglycans, prebiotics, fiber, probiotics, fatty acids, enzymes, minerals, trace elements, and vitamins. A "bioactive compound" is any compound that contributes to the health of an individual or provides a benefit to the human body beyond satisfying basic nutritional needs. The one or more additional bioactive compounds may be derived from natural sources. That is, the compounds may be obtained from extracts of plants, animals, fish, fungi, algae, or microbial fermentation. Minerals are considered to be derived from natural sources. In a preferred embodiment, the enzyme may be, for example, a protease, such as trypsin, or an enzyme extract, such as bromelain.

[0047]

[0051] In another embodiment, oleuropein and / or derivatives may be provided by any of the compositions and methods disclosed in WO 2019 / 092068 and WO 2019 / 092066, entitled "Bioconversion of oleuropein" and "Method of selecting a probiotic," respectively, and WO 2019 / 092069, entitled "Homovanilyl alcohol (HVA), HVA isomer, methods of making compositions comprising such compounds, and methods of using such compounds," each of which is incorporated herein by reference in its entirety.

[0048]

[0052] The effective amount of each of oleuropein and / or its metabolites and fisetin and / or its derivatives varies depending on the specific composition, the age and condition of the recipient, and the specific disorder or disease being treated. However, in a general embodiment, an individual can be administered 0.001 mg to 1.0 g / day, preferably 0.01 mg to 0.9 g / day, more preferably 0.1 mg to 750 mg / day, more preferably 0.5 mg to 500 mg / day, and most preferably 1.0 mg to 200 mg / day. Furthermore, the inventors have found that the active dose of oleuropein or derivatives in the combination can be reduced for equivalent efficacy.

[0049]

[0053] In some embodiments, the combination of oleuropein or metabolites and fisetin or derivatives is administered in a composition further comprising calcium. At least a portion of the calcium may be one or more calcium salts, such as calcium acetate, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium gluconate, calcium lactate, or mixtures thereof. In general embodiments, the individual is administered between 0.1 g and 1.0 g of calcium per day, preferably between 125 mg and 950 g of calcium per day, more preferably between 150 mg and 900 mg of calcium per day, more preferably between 175 mg and 850 mg of calcium per day, and most preferably between 200 mg and 800 mg of calcium per day.

[0050]

[0054] At least one oleuropein or metabolite thereof and fisetin or a derivative may be formulated in a specific ratio. In some embodiments, the formulation may include these components in the following exemplary ratios: 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:20, each of which may be fisetin:OLE in some embodiments and OLE:fisetin in other embodiments. Preferably, the ratio of OLE:fisetin is between 1:1 and 1:10.

[0051]

[0055] In an alternative embodiment, the combination of oleuropein and fisetin can be administered sequentially in a separate composition with calcium. The term "sequentially" means that calcium and at least one oleuropein or metabolite thereof are administered sequentially, such that at a first time, at least one oleuropein or metabolite thereof is administered without calcium, and at a second time (before or after the first time), calcium is administered without the combination of oleuropein and fisetin. The time between consecutive administrations can be, for example, a second or a few seconds, minutes, or hours on the same day; a day or a few days, or a few weeks on the same month; or a month or a few months on the same year.

[0052]

[0056] In some embodiments, oleuropein or a metabolite thereof and fisetin or a derivative thereof are the only polyphenols in a composition and / or the only polyphenols administered to an individual.

[0053]

[0057] The composition may comprise an effective amount of at least one oleuropein or metabolite thereof. For example, a serving or dose of the composition may comprise an effective amount, and a package may comprise one or more servings or doses. Optionally, the composition may further comprise calcium.

[0054]

[0058] The composition may include a food additive selected from the group consisting of an acidulant, a thickener, a pH adjusting buffer or pH adjusting agent, a chelating agent, a colorant, an emulsifier, an excipient, a flavoring agent, a mineral, an osmotic agent, a pharma- ceutically acceptable carrier, a preservative, a stabilizer, a sugar, a sweetener, a modifier, a vitamin, a mineral, and combinations thereof.

[0055]

[0059] The combination of oleuropein or metabolites and fisetin or derivatives can be administered in any composition suitable for human and / or animal consumption. In a preferred embodiment, such combination is administered orally or enterally (e.g., tube feeding) to an individual. For example, such combination can be administered to an individual as a beverage, food product, capsule, tablet, powder, or suspension.

[0056]

[0060] Non-limiting examples of suitable compositions include food compositions, dietary supplements, dietary supplements (e.g., liquid ONS), complete nutritional compositions, beverages, pharmaceuticals, oral dietary supplements, medical foods, nutraceuticals, Foods for Special Medical Purposes (FSMP), powdered nutritional products to be reconstituted with water or milk prior to ingestion, food additives, medicaments, drinks, pet foods, and combinations thereof.

[0057]

[0061] Food products according to the invention include, by way of example, dairy products, e.g. fermented dairy products such as yoghurt, buttermilk and the like; ice cream; condensed milk; milk, dairy cream; flavoured milk drinks; whey-based drinks; toppings; coffee creamers; chocolate; cheese-based products; soups; sauces; purees; dressings; puddings; custards; baby foods; nutritional formulas, e.g. complete nutritional formulas for infants, children, teenagers, adults, the elderly or the seriously ill; cereals and cereal bars.

[0058]

[0062] The drinks may include, for example, milk or yogurt based drinks, fermented milks, protein drinks, coffee, tea, energy drinks, soy drinks, fruit and / or vegetable drinks, fruit and / or vegetable juices.

[0059]

[0063] The combination of oleuropein or a metabolite and fisetin or a derivative may be administered in a food product further comprising an ingredient selected from the group consisting of proteins, carbohydrates, fats, and mixtures thereof.

[0060]

[0064] In one embodiment, the protein source is preferably a purified protein (i.e. isolated from the natural food source in which the protein was produced). The protein content of the composition is preferably 20-99% by weight of the composition, such as 20-90% by weight of the composition, such as 30-80% by weight of the composition, such as 40-80% by weight of the composition, such as 50-80% by weight, such as 40-70% by weight of the composition.

[0061]

[0065] Non-limiting examples of proteins or sources thereof suitable for use in the composition include hydrolyzed, partially hydrolyzed, or non-hydrolyzed proteins or protein sources. These may be derived from any known or other suitable source, such as milk (e.g., casein, whey), animal (e.g., meat, fish), grain (e.g., rice, corn), or vegetable (e.g., soy, pea). Combinations of sources or proteins may be used. Non-limiting examples of proteins or sources thereof include raw pea protein, raw pea protein isolate, raw pea protein concentrate, milk protein isolate, milk protein concentrate, casein protein isolate, casein protein concentrate, whey protein concentrate, whey protein isolate, sodium or calcium caseinate, whole milk, partially or fully skimmed milk, yogurt, soy protein isolate, and soy protein concentrate, and combinations thereof. Combinations of sources or proteins may be used. Preferred proteins include pea protein, whey protein, soy protein, and casein. Casein proteins may include, for example, sodium caseinate and calcium caseinate.

[0062]

[0066] The protein source may be provided by individual amino acids, polypeptides containing amino acids, or mixtures thereof. In many muscle growth, muscle maintenance, and / or muscle building treatments, specific amino acids that are beneficial are, for example, L-arginine, L-glutamine, lysine, and branched chain amino acids (i.e., leucine, isoleucine, and valine; particularly leucine and isoleucine). These specific amino acids may be provided as protein sources or may be added to the main source of protein. Thus, the protein source in the composition may include one or more branched chain amino acids (leucine, isoleucine, and valine), one or both of L-arginine and L-glutamine, and lysine. In a preferred embodiment, the composition includes whey protein and / or casein protein together with one or more individual amino acids, for example, one or more (or all) of leucine, isoleucine, and L-arginine.

[0063]

[0067] In one embodiment, the composition further comprises a medium chain triglyceride, such as one or more of caproic acid, caprylic acid, capric acid, and lauric acid. In one embodiment, the composition further comprises a phospholipid, such as a phosphatidylcholine.

[0064]

[0068] The composition may also contain a carbohydrate source and / or a fat source. Non-limiting examples of suitable fats include canola oil, corn oil, and high oleic sunflower oil. Non-limiting examples of suitable carbohydrates include sucrose, lactose, glucose, fructose, corn syrup solids, maltodextrin, and mixtures thereof. Additionally or alternatively, dietary fiber may be added. Dietary fiber passes through the small intestine without being enzymatically digested and acts as a natural bulking and laxative agent. Dietary fiber may be soluble or insoluble, with blends of the two generally being preferred. Non-limiting examples of suitable dietary fiber include soybean, pea, oat, pectin, guar gum, partially hydrolyzed guar gum, gum arabic, fructooligosaccharides, acid oligosaccharides, galactooligosaccharides, sialyllactose, and oligosaccharides derived from animal milk. A preferred fiber blend is a mixture of inulin and relatively short chain fructooligosaccharides. In one embodiment, the fiber content is from 2 to 40 g / L of the composition, for example from 4 to 10 g / L.

[0065]

[0069] In addition to any calcium, one or more other minerals may be used in the composition.Non-limiting examples of suitable minerals include boron, chromium, copper, iodine, iron, magnesium, manganese, molybdenum, nickel, phosphorus, potassium, selenium, silicon, tin, vanadium, zinc, or combinations thereof.

[0066]

[0070] One or more other vitamins can be used in the composition.Non-limiting examples of suitable vitamins include vitamin A, vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin or niacinamide), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine, pyridoxal, or pyridoxamine, or pyridoxine hydrochloride), vitamin B7 (biotin), vitamin B9 (folic acid), and vitamin B12 (various cobalamins; typically cyanocobalamin in vitamin supplements), vitamin C, vitamin D, vitamin E, vitamin K, folic acid, and biotin), and combinations thereof."Vitamin" includes compounds such as provitamins, their derivatives, and their analogs, which are naturally obtained from plant and animal foods or are synthesized.

[0067]

[0071] One or more food grade emulsifiers may be incorporated into the composition, such as diacetyl tartaric acid esters of mono- and diglycerides, lecithin, and / or mono- and diglycerides. Suitable salts and stabilizers may also be included.

[0068]

[0072] The compositions disclosed herein may be used in any of a variety of formulations for therapeutic administration. More specifically, the pharmaceutical compositions may include suitable pharma- ceutically acceptable carriers or diluents and may be formulated as solid, semi-solid, liquid or gaseous formulations, such as tablets, capsules, powders, granules, ointments, liquids, suppositories, injections, inhalants, gels, microspheres, and aerosols. Thus, administration of the compositions may be accomplished in a variety of ways, including oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal, and intratracheal administration. The active ingredient may be systemic after administration, or may be localized by the use of topical administration, intramural administration, or the use of an implant that acts to retain an effective dose at the implantation site.

[0069]

[0073] In pharmaceutical dosage forms, the compounds may be administered as pharma- ceutically acceptable salts. They may also be used in appropriate association with other pharmacologically active compounds. The following methods and additives are merely illustrative and in no way limiting.

[0070]

[0074] In oral formulations, the compounds can be used alone or in combination with suitable excipients for the manufacture of tablets, powders, granules or capsules, e.g., in combination with conventional excipients such as lactose, mannitol, corn starch or potato starch; in combination with binders such as microcrystalline cellulose, functional derivatives of cellulose, gum arabic, corn starch or gelatin; in combination with disintegrating agents such as corn starch, potato starch or sodium carboxymethylcellulose; in combination with lubricants such as talc or magnesium stearate; and, if desired, in combination with diluents, buffers, wetting agents, preservatives and flavoring agents.

[0071]

[0075] The composition may be administered at least 1 day per week, preferably at least 2 days per week, more preferably at least 3 or 4 days per week (e.g., every other day), most preferably at least 5 days per week, 6 days per week, or 7 days per week. The administration period may be at least 1 week, preferably at least 1 month, more preferably at least 2 months, most preferably at least 3 months, e.g., at least 4 months. In one embodiment, administration is at least daily, e.g., the subject may receive one or more doses per day. In some embodiments, administration is continued for the remainder of the individual's life. In other embodiments, administration is administered until there is no detectable symptom of the medical condition. In a specific embodiment, administration is administered until there is a detectable improvement in at least one symptom, and in further cases, administration is continued to maintain remission.

[0072] Treatment method

[0076] The compositions disclosed herein may be effective in (i) improving physiological conditions associated with metabolic fatigue in one or more cells, (ii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells, and (iii) increasing antioxidant capacity, reducing oxidative stress and / or enhancing mitochondrial function, (iv) treating or preventing calcium deficiency / depletion disorders in an individual.

[0073]

[0077] In one embodiment, at least a portion of the one or more cells is part of at least one body part selected from the group consisting of liver, kidney, brain, and skeletal muscle.

[0074]

[0078] In another embodiment, metabolic fatigue includes lack of energy, particularly physical energy, lack of vitality or reduced energy.

[0075]

[0079] In some embodiments, the methods include identifying the individual as having or at risk for the condition, prior to administration.

[0076]

[0080] In another embodiment, the disclosure provides a combination of oleuropein and / or its metabolites with fisetin and / or derivatives for treating or preventing (e.g., reducing the incidence and / or severity of) a mitochondrial-related disease or condition associated with altered mitochondrial function in an individual in need of or at risk for treatment or prevention. The method comprises orally administering an effective amount of at least one oleuropein or metabolite thereof to an individual in need of or at risk for treatment.

[0077]

[0081] Without being bound by theory, it is believed that various types of stress cause mitochondrial stress damage, which reduces the ability of mitochondria to perform many roles essential to overall cellular function.The methods disclosed herein can be useful for treating conditions involving stress damage to mitochondria.This damage can be manifested in any of a number of ways, including but not limited to mitochondrial disease.

[0078]

[0082] Mitochondrial diseases are the result of either inherited or spontaneous mutations in mitochondrial or nuclear DNA, which alter the function of proteins or RNA molecules normally present in mitochondria. However, problems in mitochondrial function may only affect certain tissues due to factors occurring during development and growth that are not yet fully understood. Even when considering tissue-specific isoforms of mitochondrial proteins, it is difficult to explain the variable patterns of organ systems affected in clinical mitochondrial disease syndromes.

[0079]

[0083] Mitochondrial diseases result from damage to mitochondria, specialized compartments present in all cells of the body except red blood cells. Mitochondria are responsible for producing over 90% of the energy the body needs to sustain life and support development. When mitochondria stop functioning, less energy is produced in the cells. The cells become damaged and even die. When this process is repeated throughout the body, the whole system begins to break down, which severely threatens the person's life. Mitochondrial diseases primarily affect children, but adult onset is also being recognized.

[0080]

[0084] Mitochondrial diseases are most likely to cause damage to cells of the brain, heart, liver, skeletal muscle, kidneys, and the endocrine and respiratory systems.

[0081]

[0085] Many symptoms of mitochondrial disorders are non-specific. Symptoms may also show a temporary course with periodic worsening. A review article on mitochondrial medicine mentions episodic migraine states, as well as muscle pain, gastrointestinal symptoms, tinnitus, depression, chronic fatigue, and diabetes, among the various manifestations of mitochondrial disease. In patients with mitochondrial disorders, clinical symptoms typically occur when energy demands are high in association with physiological stressors such as illness, hunger, excessive exercise, and extreme environmental temperatures. In addition, psychological stressors often trigger symptoms, perhaps due to the brain's energy demands being so high that the patient cannot perform sufficient ATP production.

[0082]

[0086] Depending on which cells are affected, symptoms can include loss of motor control, muscle weakness and pain, gastrointestinal problems and difficulty swallowing, poor growth, heart disease, liver disease, diabetes, respiratory complications, seizures, visual / hearing problems, lactic acidosis, developmental delays, and increased susceptibility to infections.

[0083]

[0087] Mitochondrial disorders include, but are not limited to, Alpers syndrome, Barth syndrome, beta-oxidation disorders, carnitine deficiency, carnitine-acyl-carnitine deficiency, chronic progressive external ophthalmoplegia syndrome, coenzyme Q10 deficiency, complex I deficiency, complex II deficiency, complex III deficiency, complex IV deficiency, complex V deficiency, CPT I deficiency, CPT II deficiency, creatine deficiency syndrome, cytochrome c oxidase deficiency, glutaric aciduria type 2, Kearns-Sayre syndrome, lactic acidosis, LCHAD (long-chain acyl-CoA dehydrogenase deficiency), Leber's hereditary optic neuropathy, Leigh syndrome, lethal infantile cardiomyopathy, cardiomyopathy, Luft's disease, MAD (medium-chain acyl-CoA dehydrogenase deficiency), mitochondrial cytopathy, mitochondrial DNA depletion, mitochondrial encephalomyopathy, lactic acidosis, and stroke-like symptoms, mitochondrial encephalopathy, mitochondrial myopathy, mitochondrial recessive ataxia syndrome, muscular dystrophies, myoclonic epilepsy and ragged-red fiber disease, myoneurogenic gastrointestinal encephalopathy, neuropathy, ataxia, retinitis pigmentosa, and ptosis, Pearson's syndrome, POLG mutations, pyruvate carboxylase deficiency, pyruvate dehydrogenase deficiency, SCHAD (short-chain acyl-CoA dehydrogenase deficiency), and very long-chain acyl-CoA dehydrogenase deficiency.

[0084]

[0088] Thus, one aspect of the present disclosure is a composition in unit dosage form comprising an effective amount of a combination of oleuropein and / or its metabolites and fisetin and / or derivatives for the treatment or prevention of at least a symptom selected from the group consisting of stress (e.g., early life stress and / or its effects), obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stress-related cognitive dysfunction, mood disorder (e.g., stress-induced or stress-related mood disorder), anxiety disorder (e.g., stress-induced or stress-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not due to a specific neurodegenerative disease), trauma, infection (e.g., ICU), or cancer.

[0085]

[0089] Another aspect of the present disclosure is a method of treating at least one symptom selected from the group consisting of stress, obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, cardiovascular disease, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stress-related cognitive dysfunction, mood disorder (e.g., stress-induced or stress-related mood disorder), anxiety disorder (e.g., stress-induced or stress-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not due to a specific neurodegenerative disease), trauma, infection (e.g., ICU), or cancer, in an individual having at least one symptom selected from the group consisting of stress, obesity, reduced metabolic rate, metabolic syndrome, diabetes mellitus, cardiovascular disease, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stress-related cognitive dysfunction, mood disorder (e.g., stress-induced or stress-related mood disorder), anxiety disorder (e.g., stress-induced or stress-related anxiety disorder), and age-related neuronal cell death or dysfunction (e.g., age-related neuronal cell death or dysfunction not due to a specific neurodegenerative disease), trauma, infection (e.g., ICU), or cancer.

[0086]

[0090] In one embodiment of these methods, the hyperlipidemia treated or prevented comprises hypertriglyceridemia. In one embodiment of these methods, the hyperlipidemia treated or prevented comprises an increase in free fatty acids. In one embodiment of these methods, the age-related neuronal cell death or dysfunction treated or prevented is by administration of the composition to a middle-aged or elderly person, such as an elderly person.

[0087]

[0091] The stress to be treated or prevented can be early life stress, i.e. stress experienced during the period from birth to 5 years of age. Early life stress has been reported to have a significant adverse effect on cognitive performance, including psychological parameters such as increased incidence or susceptibility to depression, anxiety, and abnormal risk-taking behavior. Higher incidence of attention deficit / hyperactivity disorder (ADHD), post-traumatic stress disorder (PTSD), and major depression has been reported in individuals who have experienced early life stress.

[0088]

[0092] Another aspect of the present disclosure is a method of delaying the onset of metabolic decline, maintaining muscle mass, reducing oxidative stress, maintaining immune function, and / or maintaining cognitive function in healthy middle-aged and older adults.

[0089]

[0093] The compositions disclosed herein can also be used to treat any of a variety of additional diseases and conditions in which defective or reduced mitochondrial activity is involved in the pathophysiology of the disease or condition, or in which increased mitochondrial function produces the desired beneficial effect.Non-limiting examples of such conditions include male infertility associated with reduced sperm motility, macular degeneration, and other age-related and genetic eye disorders and hearing loss (e.g., age-related hearing loss).

[0090]

[0094] Yet another aspect of the present disclosure is a unit dosage form comprising a combination of oleuropein and / or derivatives thereof and fisetin and / or derivatives thereof in an effective amount for at least one of: i) treating, reducing the incidence, or reducing the severity of a mitochondrial-related disease or condition associated with altered mitochondrial function; (ii) improving a physiological condition associated with metabolic fatigue in one or more cells; (iii) increasing mitochondrial energy and mitochondrial calcium uptake in one or more cells; and (iv) treating or preventing calcium deficiency / depletion disorders; (v) increasing metabolic rate; (vi) improving or maintaining cognitive function; and (vii) enhancing or maintaining mitochondrial function.

[0091]

[0095] In one embodiment, physiological conditions associated with metabolic fatigue include muscle fatigue or weakness, lack of energy, lack of physical energy, lack of vitality or reduced energy.

[0092]

[0096] In one embodiment, the present invention relates to a method of preventing (e.g., reducing the incidence and / or severity) or treating exercise-induced muscle fatigue, in particular reducing muscle fatigue, in an individual participating in exercise comprising at least one of 1) resistance exercise, 2) anaerobic exercise or repeated sprint type exercise, or 3) endurance exercise, comprising orally administering to the individual an effective amount of a combination of fisetin or a derivative and at least one oleuropein or metabolite thereof. In some embodiments, the method comprises identifying the individual as having or at risk for a condition prior to administration.

[0093]

[0097] In one embodiment, the composition is administered in at least one dose during at least one time period selected from the group consisting of: (i) a pre-exercise time between 1 hour and 1 second before the start of exercise, e.g., between 30 minutes and 1 minute before the start of exercise; (ii) an exercise time from the start of exercise to the end of exercise; and (i) a post-exercise time between 1 second after the end of exercise and 1 hour after the end of exercise, e.g., between 1 minute after the end of exercise and 30 minutes after the end of exercise.

[0094]

[0098] In further embodiments, the unit dosage form consists essentially of oleuropein and / or its metabolites in combination with fisetin and / or derivatives, in some embodiments, one or more of these compounds may be isolated compounds.

[0095]

[0099] The disclosure also provides a kit comprising a combination of oleuropein and / or its metabolites in combination with fisetin and / or derivatives in one or more containers. In one embodiment of the kit, the one or more containers include at least one first container that contains oleuropein and / or metabolites separately from fisetin and / or derivatives contained in at least one second container, and the kit further comprises instructions for combining the oleuropein with fisetin to form a unit dosage form.

[0096]

[0100] In one embodiment of the kit, the combination may be provided together as one or more prepackaged unit dosage forms, e.g., as separate containers each containing a dry powder, such that each container contains one prepackaged unit dosage form.

[0097]

[0101] In another embodiment, the kit can include multiple compositions to be mixed together to form one or more of the compositions disclosed herein.For example, the kit can contain two or more dry powders in separate containers that are related to each other, each of which contains a portion of the final unit dosage form.As a non-limiting example of such an embodiment, the kit can include one or more first containers that contain oleuropein, and also one or more second containers that contain fisetin.The contents of one of the first containers can be mixed with one of the second containers to form at least a portion of the unit dosage form of the composition.

[0098]

[0102] The above dosing examples do not require uninterrupted daily administration. Instead, there may be several short interruptions in administration, such as a 2-4 day break during the administration period. The ideal duration of administration of the compositions of the present invention can be determined by one of skill in the art.

[0103] EXAMPLES

[0099]

[0104] The following non-limiting examples provide experimental data supporting the compositions and methods disclosed herein.

[0100]

[0105] Example 1

[0106] To test the effect of oleuropein (or oleuropein aglycone), fisetin and their combination in living cells, we measured mitochondrial calcium elevation in HeLa cells. HeLa cells were purchased from ATCC. HeLa cells were seeded in 96-well plates at a density of 50,000 cells / well in Minimum Essential Medium (DMEM, Gibco), high glucose, +10% fetal bovine serum. Mitochondrial calcium measurements were performed using HeLa cells (Sirion biotech) infected with an adenovirus expressing mitochondrial modified aequorin, a calcium sensor targeted in mitochondria (Montero et al., 2004). For aequorin reconstitution, 24 hours after infection, standard medium (145 mM NaCl, 5 mM KCl, 1 mM MgCl) supplemented with 1 μM wild-type coelenterazine was used. 2 , 1 mM CaCl 2 Cells were incubated in 10 mM NaCl, 10 mM glucose, and 10 mM Hepes, pH 7.4 for 2 hours at room temperature (22±°C). For treatment, compounds were added directly to cell or myotube cultures 2 hours before measurement. Luminescence was measured with a FLIPR Tetra Aequorin (Molecular Devices). Mitochondrial calcium elevation was obtained by stimulating cells with 100 μM histamine. Conversion of luminescence data to calcium concentration (Calibration) was performed using an algorithm as previously described (Alvarez & Montero, 2002). Quantification was performed using a custom module analysis based on Excel (Microsoft) and GhaphPad Prism 7.02 (GhaphPad) software.

[0101]

[0107] As shown in Figure 2, the effect of the combination of oleuropein and fisetin is greater than the effect of fisetin or oleuropein alone on activating mitochondria by increasing mitochondrial Ca2+ in HeLa cells, and Figure 3 shows that fisetin synergizes with oleuropein to activate mitochondria by increasing mitochondrial Ca2+ in HeLa cells.

[0102]

[0108] It should be understood that various changes and modifications to the presently preferred embodiments described herein will be apparent to those skilled in the art. Such changes and modifications can be made without departing from the spirit and scope of the present subject matter and without diminishing its intended advantages. Accordingly, such changes and modifications are intended to be encompassed by the appended claims.

Claims

1. A composition comprising a therapeutically effective amount of oleuropein and / or its metabolites and fisetin and / or derivatives for use in the treatment or prevention of calcium deficiency / depletion disorders in an organism, comprising: (i) improvement of physiological conditions associated with metabolic fatigue in one or more cells; (ii) increased mitochondrial energy and mitochondrial calcium uptake in one or more cells; and (iii) increased antioxidant capacity, reduced oxidative stress and / or enhanced mitochondrial function; and (iv) a combination of oleuropein and / or its metabolites and fisetin and / or derivatives.

2. The composition according to claim 1, wherein at least a portion of the one or more cells is part of at least one body part selected from the group consisting of the liver, kidney, brain, and skeletal muscle.

3. The composition according to claim 1, wherein the physiological state related to metabolic fatigue includes muscle fatigue or weakness, lack of energy, lack of physical energy, lack of vitality or decreased vitality.

4. The composition according to claim 1 for preventing or treating muscle fatigue caused by exercise.

5. The composition according to claim 1, comprising administering, orally, a therapeutically effective amount of oleuropein and / or its metabolites in combination with fisetin and / or derivatives on a daily basis for at least one week.

6. The composition according to claim 1, wherein the metabolite of oleuropein is selected from the group consisting of oleuropein aglycone, hydroxytyrosol, elenolic acid, homovanillyl alcohol, isohomovannillyl alcohol, glucuronide conjugates thereof, sulfate forms thereof, derivatives thereof, and mixtures thereof.

7. The composition according to claim 1, wherein the derivative of fisetin is selected from the group consisting of its glucuronidated form, its sulfate form, its derivatives, and mixtures thereof.

8. The composition according to claim 1, further comprising at least one compound selected from the group consisting of antioxidants, anti-inflammatory compounds, glycosaminoglycans, prebiotics, fiber, probiotics, fatty acids, enzymes, minerals, trace elements and / or vitamins.

9. The composition according to claim 1, selected from the group consisting of food compositions, nutritional supplements, nutritional compositions, oral nutritional supplements, medical foods, nutraceuticals, beverages, powdered nutritional products that are reconstituted with water or milk before ingestion, food additives, foods for special medical purposes (FSMPs), pharmaceuticals, drinks, pet food, and combinations thereof.

10. The composition according to claim 1, which is in the form of a solid powder, powder stick, capsule, or solution.

11. The composition according to claim 1, wherein the combination of oleuropein and / or its metabolites and fisetin and / or derivatives in a therapeutically effective amount is administered in a food product further comprising components selected from the group consisting of proteins, carbohydrates, fats, and mixtures thereof.

12. A composition comprising a therapeutically effective dose of oleuropein and / or its metabolites and fisetin and / or derivatives for treating, reducing the incidence, and / or reducing the severity of mitochondrial-related diseases or conditions associated with altered mitochondrial function in individuals who require or are at risk thereof, The aforementioned composition is a composition intended for oral administration.

13. The composition according to claim 12, wherein the mitochondrial-related disease or condition is selected from the group consisting of stress, physiological aging, obesity, decreased metabolic rate, metabolic syndrome, diabetes mellitus, complications of diabetes, hyperlipidemia, neurodegenerative disease, cognitive impairment, stress-induced or stress-related cognitive impairment, mood disorders, anxiety disorders, age-related neuronal cell death or dysfunction, musculoskeletal disorders, frailty, pre-frailty, chronic kidney disease, renal failure, trauma, infection, cancer, hearing loss, macular degeneration, myopathy and dystrophy, and combinations thereof.

14. A composition comprising a therapeutically effective dose of oleuropein and / or its metabolites and fisetin and / or derivatives for the purpose of delaying the onset of metabolic decline, maintaining muscle mass and / or muscle function, reducing oxidative stress, maintaining immune function, and / or maintaining cognitive function in healthy middle-aged and elderly individuals.

15. A composition comprising a therapeutically effective amount of oleuropein and / or its metabolites and fisetin and / or derivatives for enhancing at least one of mental or muscular performance in an individual.

16. A composition comprising a therapeutically effective amount of oleuropein and / or its metabolites and fisetin and / or derivatives for improving or maintaining cognitive function in an individual.

17. The composition according to claim 16, wherein the cognitive function is selected from the group consisting of perception, memory, attention, speech comprehension, speech production, reading comprehension, image creation, learning, reasoning, and combinations thereof.

18. The composition according to claim 1, wherein the combination of oleuropein and / or its metabolites and fisetin and / or derivatives in a therapeutically effective amount is administered in a composition further containing calcium.

19. The composition according to claim 1, wherein the individual is a middle-aged or elderly person, an elderly person, or an ICU patient.

20. A unit dosage form comprising, in an effective amount, a combination of oleuropein and / or its derivatives and fisetin and / or its derivatives for at least one of the following: (i) treatment of, reduction of incidence of, or reduction of severity of mitochondrial-related diseases or conditions associated with altered mitochondrial function; (ii) improvement of physiological conditions associated with metabolic fatigue in one or more cells; (iii) increase of mitochondrial energy and mitochondrial calcium uptake in one or more cells; and (iv) treatment or prevention of calcium deficiency / depletion disorders; (v) increase of metabolic rate; (vi) improvement or maintenance of cognitive function; or (vii) improvement or maintenance of mitochondrial function.

21. The unit dosage form according to claim 20, wherein the physiological state related to metabolic fatigue includes muscle fatigue or weakness, lack of energy, lack of physical energy, lack of vitality or decreased vitality.

22. The unit dosage form according to claim 20 or 21, which essentially consists of a combination of oleuropein and / or its metabolites and fisetin and / or derivatives.

23. A kit comprising one or more containers containing a combination of oleuropein and / or its metabolites and fisetin and / or derivatives.

24. The kit according to claim 23, wherein the one or more containers include at least one first container which contains the oleuropein and / or metabolites separately from the fisetin and / or derivatives contained in at least one second container, and the kit further includes instructions for mixing the oleuropein with fisetin to form a unit dosage form.

25. The kit according to claim 23 or 24, wherein each of the one or more containers contains a unit dosage form of a combination of oleuropein and / or its metabolite and fisetin and / or a derivative.

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