Compositions for increasing spermidine production of microbiota

Abstract

The present invention relates to a composition comprising at least one protein source and at least one prebiotic source for use in increasing spermidine production in the gut microbiota. The compositions are useful for the prevention and / or treatment of age-related disorders.

Description

Technical Field

[0001] This invention relates to compositions comprising at least one protein source and at least one prebiotic source, wherein the compositions are used to increase spermidine production in the gut microbiota. The compositions may be used to prevent and / or treat age-related disorders. Background Technology

[0002] Polyamines are small polycationic molecules with a wide range of biological functions, including gene regulation, stress resistance, cell proliferation and differentiation, and are associated with both eukaryotic and prokaryotic cells.

[0003] Spermine, putrescine, and spermine are the major polyamines in eukaryotes. These polyamines regulate important cellular functions, including growth and proliferation, RNA and DNA stability, RNA-to-protein translation, autophagy, and immune responses.

[0004] Spermine has emerged as an anti-aging metabolite, crucial for cellular and tissue function in various organisms, including humans. Several sets of evidence support its anti-aging properties: during aging, spermine levels decrease in most studied tissues and species. 1 Spermine supplementation has extended the lifespan and bodily functions of multiple species. 1 Epidemiological data support the view that increased dietary intake reduces, delays, or interrupts age-related deterioration, including cancer, cardiovascular disease, and cognitive impairment. 2-4 Damaged intestinal barrier integrity 5-6 Leaky gut or a damaged gut microbiome are contributing factors to age-related inflammation. 7 .

[0005] Ingested foods (such as soybeans, mushrooms, wheat germ, beef, pork, chicken liver, oranges, horned snail viscera, and green tea leaves) are the main direct sources of spermidine in the cavity.

[0006] However, the upper part of the intestine absorbs most of these compounds. Therefore, direct administration of spermidine sources has a limited effect on circulating spermidine levels. 8 .

[0007] The gut microbiota is believed to be primarily responsible for spermidine levels in the lower intestine. Gut bacteria synthesize polyamines using arginine, ornithine, or lysine as substrates. Polyamines (including spermidine) produced in the colonic lumen are translocated into the bloodstream via the colonic mucosa, where they may affect certain cellular functions in tissue- or cell-specific ways, thus leading to anti-aging effects.

[0008] On average, 3g to 18g of food escapes digestion and absorption in the small intestine and enters the colon. Bacteria in the colon can further break them down into small peptides and free amino acids, which can be further metabolized into compounds such as polyamines (including spermidine).

[0009] However, there are no studies comparing the effects of different dietary nutrients on the production of spermidine by microorganisms.

[0010] Therefore, there is clearly an unmet need for new compositions that increase spermidine production in the gut microbiota and, as a result, increase resistance to age-related symptoms, improve physiological states or disorders associated with cellular senescence, improve physiological states associated with metabolic fatigue in one or more cells, increase mitochondrial energy in one or more cells, increase antioxidant capacity, reduce oxidative stress and / or enhance mitochondrial function, improve subject activity and / or improve health and / or lifespan.

[0011] References

[0012] 1.Hofer, SJ; Simon, AK; Bergmann, M.; Eisenberg, T.; Kroemer, G.; Madeo, F., Mechanisms of spermidine-induced autophagy and geroprotection. Nature Aging 2022, 2 (12), 1112-1129.

[0013] 2.Kiechl, S.; Pechlaner, R.; Willeit, P.; Notdurfter, M.; Paulweber,B.; Willeit, K.; Werner, P.; Ruckenstuhl, C.; Iglseder, B.; Weger, S.;Mairhofer, B.; Gartner, M.; Kedenko, L.; Chmelikova, M.; Stekovic, S.;Stuppner, H.; Oberhollenzer, F.; Kroemer, G.; Mayr, M.; Eisenberg, T.; Tilg,H.; Madeo, F.; Willeit, J., Higher spermidine intake is linked to lowermortality: a prospective population-based study. The American Journal ofClinical Nutrition 2018, 108 (2), 371-380.

[0014] 3.Schroeder, S.; Hofer, SJ; Zimmermann, A.; Pechlaner, R.;Dammbrueck, J.; Pendle, T.; Marcello, GM; Pogatschnigg, V.; Bergmann, M.; Müller, M.; Gschiel, V.; Ristic, S.; Tadic, J.; Iwata, K.; Richter, G.; Farzi,A.; Uchal, M.; Schäfer, U.; Poglitsch, M.; Royer, P.; Mekis, R.; Agreiter, M.;Tölle, RC; Sotonyi, P.; Willeit, J.; Mairhofer, B.; Niederkofler, H.;Pallhuber, I.; Runger, G.; Tilg, H.; Defrancesco, M.; Marksteiner, J.;Sinner, F.; Magnes, C.; Pieber, T.R.; Holzer, P.; Kroemer, G.; Carmona-Gutierrez, D.; Scorrano, L.; Dengjel, J.; Madl, T.; Sedej, S.; Sigrist, SJ; Rácz, B.; Keechle, S.; Eisenberg, T.; Madeo, F., Dietary spermidineimproves cognitive function. Cell Reports 2021, 35 (2) 108985.

[0015] 4. Wu, H.; Wang, J.; Jiang, H.; Liu, X.; Sun, X.; Chen, Y.; Hu, C.; Wang, Z.; Han, T.; Sun, C.; Wei, W.; Jiang, W., The association of dietary spermidine with all-cause mortality and CVD mortality: The U.S. National Health and Nutrition Examination Survey, 2003 to 2014. Frontiers in Public Health 2022, 10:949170.

[0016] 5. Ma L, Ni Y, Wang Z, Tu W, Ni L, Zhuge F, Zheng A, Hu L, Zhao Y, Zheng L, Fu Z. Spermidine improves gut barrier integrity and gut microbiota function in diet-induced obese mice. Gut Microbes. November 9, 2020; 12(1):1 - 19. doi: 10.1080 / 19490976.2020.1832857. PMID: 33151120; PMCID: PMC7668533.

[0017] 6. Jacopo J.V. Branca, Massimo Gulisano, Claudio Nicoletti, Intestinal epithelial barrier functions in ageing, Ageing Research Reviews, Volume 54, 2019, 100938, ISSN 1568 - 1637.

[0018] 7.Lan Zhang, Junbin Yan, Chi Zhang, Shuyan Feng, Zheli Zhan, YangBao, Shuo Zhang, Guanqun Chao, Improving intestinal inflammaging to delayaging? A new perspective, Mechanisms of Ageing and Development, Volume 214, 2023, 111841, ISSN 0047-6374.8. Senekowitsch, S.; Wietkamp, ​​E.; Grimm, M.;Schmelter, F.; Schick, P.; Kordowski, A.; Sina, C.; Otzen, H.; Weitschies, W.; Smollich, M., High-Dose Spermidine Supplementation Does Not IncreaseSpermidine Levels in Blood Plasma and Saliva of Healthy Adults: A RandomizedPlacebo-Controlled Pharmacokinetic and Metabolomic Study. Nutrients 2023, 15(8), 1852. Summary of the Invention

[0019] The present invention provides compositions that increase spermidine production in the gut microbiota.

[0020] Specifically, the present invention provides a composition comprising at least one protein source and at least one prebiotic source, the composition being used to increase spermidine production in the gut microbiota.

[0021] In one embodiment, the composition comprises a protein source selected from at least one of the following: legumes, grains, nuts, algae, oil crops and / or their cakes, animal proteins, or combinations thereof.

[0022] In one implementation, the legume is selected from at least one of the group consisting of: broad beans, chickpeas, green beans, and lentils.

[0023] In one implementation, the cereal is selected from at least one of the group consisting of: rice, oats, quinoa, wheat, and barley.

[0024] In one implementation, the nut is selected from at least one of walnuts, almonds, and pistachios.

[0025] In one implementation, the algae is selected from at least one of seaweed and microalgae.

[0026] In one implementation, the oilseed crop and / or its cake is selected from soybeans, rapeseed, peanuts, and bamboo.

[0027] In one implementation, the animal protein is selected from at least one of the following: milk, eggs, seafood, poultry, red meat, and white meat.

[0028] In one embodiment, the composition comprises a prebiotic source, which is at least one oligosaccharide.

[0029] In one embodiment, the composition comprises at least one oligosaccharide selected from the group consisting of: fructooligosaccharides, inulin, guar gum, galactooligosaccharides, β-glucan, resistant starch, arabinoxylan, pectin, raffinose, polydextrose, agar, alginate, or combinations thereof.

[0030] In one embodiment, the composition comprises an effective amount of at least one protein source and at least one prebiotic source.

[0031] In one embodiment, the composition comprises a protein source administered to the subject in an amount of at least about 1 mg to at least about 2400 mg per kg of the subject's body weight.

[0032] In one embodiment, the composition comprises a prebiotic source administered to the subject in an amount of at least about 10 mg to at least about 1000 mg per kg of subject body weight.

[0033] In one embodiment, the composition comprises at least one protein source (which is fava bean) and at least one prebiotic source (which is fructooligosaccharide).

[0034] In one embodiment, the composition comprises:

[0035] - A minimum of about 1 mg to a minimum of about 2400 mg of fava bean protein per kg of the subject's body weight.

[0036] - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

[0037] In one embodiment, the composition comprises at least one protein source (which is legume protein) and at least one prebiotic source (which is fructooligosaccharide).

[0038] In one embodiment, the composition comprises:

[0039] - A minimum of about 1 mg to a minimum of about 2400 mg of legume protein per kg of the subject's body weight.

[0040] - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

[0041] In one embodiment, the composition is used to prevent and / or treat age-related disorders.

[0042] In one implementation, the age-related impairment is selected from at least one of the following groups: physiological states or impairments related to cellular senescence, physiological states related to metabolic fatigue in one or more cells, excessive catabolism, oxidative stress, reduced antioxidant capacity and / or mitochondrial function, impaired activity, healthy period and / or lifespan of the subject, muscle dysfunction, including sarcopenia, weakness, inclusion body myositis, myopathy / rhabdomyolysis induced by drugs such as corticosteroids or statins, muscle wasting induced by inactivity or hospitalization, myasthenia gravis, immune dysfunction, cognitive impairment, including delirium, dementia, learning disabilities, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD), cardiovascular disease, impaired intestinal barrier integrity or leaky gut, impaired gut microbiome, and age-related inflammation.

[0043] In one embodiment, the composition is used to: (i) increase resistance to age-related symptoms in a subject; (ii) improve physiological states or disorders associated with cellular senescence; (iii) improve physiological states associated with metabolic fatigue in one or more cells; (iv) increase mitochondrial energy in one or more cells; (v) increase antioxidant capacity, reduce oxidative stress, and / or enhance mitochondrial function; (vi) improve activity and / or (vii) improve the subject's health and / or lifespan.

[0044] In one embodiment, the composition is in the form of a food composition, dietary supplement, nutritional composition, nutritional product, powdered nutritional product reconstituted in water or milk before consumption, food additive, medicine, beverage, and drink, including coffee-based products, oral nutritional supplements (ONS), complete nutritional formulas, pharmaceuticals, medical or food products.

[0045] In one implementation scheme, the subjects are humans, livestock, or pets.

[0046] Another aspect of this disclosure is the use of the composition in subjects who require at least one of the following: (i) increased resistance to age-related conditions; (ii) improved physiological states or disorders associated with cellular senescence; (iii) improved physiological states associated with metabolic fatigue in one or more cells; (iv) increased mitochondrial energy in one or more cells; (v) increased antioxidant capacity, reduced oxidative stress, and / or enhanced mitochondrial function; (vi) improved activity and / or (vii) improved individual health and / or lifespan.

[0047] The benefit of this improvement is the prevention and / or treatment of age-related disorders.

[0048] The benefits of such improvements also include: (i) increased resistance to age-related symptoms; (ii) improved physiological states or disorders associated with cellular senescence; (iii) improved physiological states associated with metabolic fatigue in one or more cells; (iv) increased mitochondrial energy in one or more cells; (v) increased antioxidant capacity, reduced oxidative stress and / or enhanced mitochondrial function; (vi) improved activity and / or (vii) improved health and / or lifespan of subjects.

[0049] Other features and advantages are described herein and will be apparent from the following figures and detailed description. Attached Figure Description

[0050] Figure 1 Using the in vitro fermentation system from Example 1, the protein source and prebiotic FOS produced by the microorganisms Effects of spermine .

[0051] Proteins were digested according to the INFogEST protocol, and small peptides and free amino acids were removed by dialysis using a 14 kDa molecular weight cutoff. In vitro fermentation was performed on three fecal samples, and spermidine concentrations at 24 hours are shown in (A). FOS was added to the fermentation medium, and the results are shown in (B). Data are mean ± SEM. Statistical analysis was performed using one-way ANOVA, followed by Tukey post-hoc analysis. No shared letters were used between groups to indicate statistically significant differences.

[0052] Figure 2 Effects of FOS on the production of spermidine (A) and putrescine (B) by the gut microbiota in Example 1 .

[0053] Experimental methods followed the methods and materials described in Example 1 of the embodiments. The in vitro system was assembled in the upper and lower intestines. Data are mean ± sem, n=3. The effect of FOS on each protein source was tested by Student's t-test, **p<0.01, *p<0.05, §p<0.1 (p<0.05 in post-hoc analysis).

[0054] Figure 3 Using in vitro fermentation experiments from Example 1, in time-course studies, arginine in the protein and The relationship between spermidine production and .

[0055] Experimental conditions are shown in the methods and materials of Example 1 in the examples. The amount of arginine was obtained from the manufacturer's analytical report or production instructions. Fermentation medium was collected at 6 h, 24 h, and 48 h for polyamine analysis. Linear regression analysis was performed at 6 h (A), 24 h (B), and 48 h (C), and the p-values ​​and R² are shown below each graph.

[0056] Figure 4 Microbial production of spermidine (A) and putrescine (B) from the in vitro fermentation experiment in Example 2 .

[0057] Data are the mean + / - SEM values ​​from experiments with three different microbial communities. Data were normalized to the amount of enzyme only and expressed as a ratio to enzyme only. Results were analyzed using a one-way ANOVA followed by a Tukey post-hoc analysis. No shared letter between groups indicated statistical significance (p < 0.05). Detailed Implementation

[0058] definition

[0059] Some definitions are provided below. However, definitions may be located in the “Implementation Scheme” section below, and the title “Definitions” above does not mean that such disclosures in the “Implementation Scheme” section are not definitions.

[0060] As used in this disclosure and the appended claims, the singular forms “a,” “an,” and “the” include plural references unless the context clearly specifies otherwise. Thus, for example, references to “a vitamin” or “the vitamin” cover embodiments having a single vitamin and embodiments having two or more vitamins.

[0061] The term "including / comprises" will be interpreted as inclusive rather than exclusive. Similarly, the terms "including / comprises" and "or" should be considered inclusive unless the context explicitly prohibits this interpretation. However, the compositions disclosed herein may not contain any elements not specifically disclosed herein. Therefore, the disclosure of embodiments using the term "including / comprises" includes both embodiments that "consist substantially of the specified components" and embodiments that "consist of the specified components."

[0062] The terms “at least one of X or Y” and “and / or” used in the appropriate contexts of “at least one of X or Y” and “X and / or Y” should be interpreted as “X” or “Y” or “X and Y”. For example, “at least one of resistance or recovery” and “resistance and / or recovery” should be interpreted as “resistance” or “recovery” or “both resistance and recovery”.

[0063] In its use herein, the terms “example” and “such as” (especially when followed by a list of terms) are exemplary and illustrative only and should not be considered exclusive or comprehensive. As used herein, “associated” or “related” to another condition means that the two conditions occur simultaneously, preferably means that the two conditions are caused by the same underlying condition, and most preferably means that one of the identified conditions is caused by the other identified condition.

[0064] The “subject” is a mammal, preferably a human, livestock, or pet. Specifically, the term “livestock” may include, but is not limited to, horses (e.g., pets or horses undergoing medical treatment), or cattle or poultry (e.g., cattle or poultry used in agriculture). In particular, the term “pet” means any animal that can benefit from or enjoy the compositions provided in this disclosure. For example, a pet can be a bird, a bovine, a canine, a equine, a feline, a goat, a wolf, a rodent, a sheep, or a pig, but a pet can also be any suitable animal.

[0065] "Prevention" includes reducing the risk, incidence, and / or severity of a condition or disorder. The term "treatment" includes both preventative or averting treatment (preventing and / or delaying the development of a target pathological condition or disorder) and curative, therapeutic, or disease-modifying treatment, including therapeutic measures that cure, delay, alleviate symptoms of a diagnosed pathological condition or disorder, and / or halt its progression; and treatment of patients at risk of or suspected of having the disease, as well as treatment of patients who are ill or have been diagnosed with a disease or medical condition.

[0066] The term "treatment" does not necessarily mean that a subject is treated until fully recovered. The term "treatment" also refers to health maintenance and / or promotion in individuals who do not have a disease but may be susceptible to unhealthy conditions. The term "treatment" is also intended to include intensifying or otherwise enhancing one or more major preventative or therapeutic measures. As a non-limiting example, treatment can be administered by a patient, caregiver, physician, nurse, or other healthcare professional.

[0067] As used in this disclosure and the appended claims, the term "upper gastrointestinal tract" refers to the gastrointestinal tract including the small intestine, oral cavity, pharynx, esophagus, stomach, and duodenum.

[0068] As used in this disclosure and the appended claims, the term "lower gastrointestinal tract" refers to the gastrointestinal tract that includes the remainder of the small intestine (i.e., the jejunum and ileum) and the entirety of the large intestine (i.e., the cecum, colon, rectum, and anal canal). Bacteria can be found in the gastrointestinal tract, particularly in the intestine.

[0069] As used in this article, the “effective amount” for prevention or treatment is the amount used to prevent an individual’s defect, treat their disease or medical condition, or more generally, the amount used to alleviate an individual’s symptoms, manage the progression of their disease, or provide them with nutritional, physiological, or medical benefits.

[0070] As used herein, the terms “food,” “food product,” and “food composition” mean a product or composition intended for oral ingestion by humans or other mammals and comprising at least one nutrient for humans or other mammals.

[0071] As used herein, “nutritional composition” and “nutritional product” include any number of food ingredients and optional additional ingredients that may be necessary for product function and fully comply with all applicable regulations. It can refer to liquids, powders, gels, pastes, solids, concentrates, suspensions, or ready-to-use enteral formulations, oral formulations, children's formulations, adult formulations, porridges and / or cereals, food products, food compositions, infant food, and pet food. Optional ingredients may include, but are not limited to, common food additives such as one or more acidifiers, additional thickeners, buffers or pH adjusters, chelating agents, colorants, emulsifiers, excipients, flavoring agents, minerals, penetrants, pharmaceutically acceptable carriers, preservatives, stabilizers, sugars, sweeteners, texture agents, and / or vitamins. Optional ingredients may be added in any suitable amount.

[0072] "Prebiotics" generally refer to indigestible food components that selectively stimulate the growth and / or activity of microorganisms present in the host's gut to have a beneficial effect on the host, thereby attempting to improve the host's health.

[0073] The term "oligosaccharide" refers to carbohydrates with a degree of polymerization (DP) ranging from 2 to 20 (inclusive), but excluding lactose. In some embodiments of the invention, the DP of the carbohydrate is in the range of 3 to 20.

[0074] The terms "galacto-oligosaccharide" and "GOS" are used interchangeably. They refer to oligosaccharides that contain two or more galactose molecules, are uncharged, and do not have N-acetyl residues (i.e., they are neutral oligosaccharides).

[0075] The terms "fructooligosaccharide" and "FOS" are used interchangeably. They refer to oligosaccharides that contain short fructose chains.

[0076] The term "guar gum" refers to a galactomannan polysaccharide composed of galactose and mannose.

[0077] The term "β-glucan" refers to a polysaccharide composed of β-D-glucose.

[0078] The term "resistant starch" refers to a polysaccharide composed of amylose and amylopectin.

[0079] The term "arabinoxylan" refers to hemicellulose composed of arabinose and xylose.

[0080] The term "pectin" refers to a polysaccharide mainly composed of galacturonic acid and other sugar residues with lower abundance, such as xylose, apigenin, rhamnose, galactose, and arabinose.

[0081] The term "raffinose" refers to a trisaccharide composed of galactose, glucose, and fructose.

[0082] The term "polydextrose" refers to a synthetic polysaccharide composed of glucose.

[0083] The term "agar" refers to a polysaccharide composed of agarose and agar gum.

[0084] The term "alginate" refers to a polysaccharide composed of β-D-mannuronic acid and α-L-guluronic acid.

[0085] The terms "oligosaccharide" or "oligosaccharide mixture" are used interchangeably. In some advantageous embodiments, the oligosaccharides in the oligosaccharide mixture are milk oligosaccharides, milk-derived oligosaccharides, or milk-derived oligosaccharides (all abbreviated as "BMO").

[0086] As used herein, “cognitive function” refers to any mental process involving symbolic operations, such as perception, memory, attention, verbal comprehension, verbal production, reading comprehension, image creation, learning, and reasoning, preferably at least memory. Methods for measuring cognitive function are well-known and may include, for example, individual tests or battery tests targeting any aspect of cognitive function. One such test is the Prudhoe Cognitive Function Test by Margallo-Lana et al., published in (2003) J. Intellect. Disability Res. 47:488-492. Another such test is the Mini Mental State Exam (MMSE), which is designed to assess orientation to time and place, immediate recall, attention and calculation, delayed recall, language use and comprehension, and repetitive and complex commands. As used herein, “cognitive impairment” refers to any condition that impairs cognitive function. Non-limiting examples of cognitive impairment include delirium, dementia, learning disabilities, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD). "Stress-induced or stress-related cognitive impairment" refers to cognitive dysfunction that is induced or associated with stress.

[0087] As used herein, the term "unit dosage form" refers to a physically discrete unit suitable for use as a unit dose in human and animal subjects, each unit containing a predetermined amount of the composition disclosed herein, associated with a pharmaceutically acceptable diluent, carrier, or mediator, sufficient to produce the desired effect. The specifications of a unit dosage form depend on the specific compound used, the effect to be achieved, and the pharmacodynamics associated with each compound in the host.

[0088] The “subject” or “individual” of this invention is a human adult subject, preferably a healthy adult, animal, or pet who requires improvement in motivation and / or mental performance by modulating glutathione levels in the brain. The compositions of this invention can be advantageously used to increase glutathione levels in the brain for the prevention or treatment of conditions or diseases characterized by low glutathione levels in the brain, whether temporary or chronic.

[0089] Implementation Plan

[0090] The inventors have surprisingly discovered that a combination of proteins and prebiotics from specific sources promotes spermidine production in the gut microbiota.

[0091] The inventors tested the concept of dietary protein as a source of microbial polyamine production, particularly spermidine, using an in vitro digestion and fermentation platform. Example 1, conducted via ProDigest, demonstrates that protein alone is insufficient to stimulate the gut microbiota to produce spermidine. The microbiota requires a carbon source such as FOS to produce spermidine. More importantly, an effect of FOS on spermidine was observed, but not on putrescine, indicating that some, but not all, microbial polyamine synthesis pathways are sensitive to prebiotic fermentation. Furthermore, the addition of the carbon source FOS extended microbial spermidine production by more than 6 hours. This further highlights the crosstalk between polyamine synthesis and fiber fermentation in the gut ecosystem.

[0092] In Example 1, the inventors found that fava bean protein was highly effective in increasing spermidine compared to the control group, where the concentration of putrescine was similar in all groups. The fava bean protein concentrate tested in this Example 1 contained 40% (w / w) non-protein material.

[0093] Therefore, one aspect of this disclosure is a composition comprising at least one protein source and at least one prebiotic source, the composition being used to increase spermidine production in the gut microbiota of a subject.

[0094] In one embodiment, the composition comprises a protein source selected from at least one of the following: legumes, grains, nuts, algae, oil crops and / or their cakes, animal proteins, or combinations thereof.

[0095] In one implementation, the legume is selected from at least one of the group consisting of: broad beans, chickpeas, green beans, and lentils.

[0096] In one implementation, the cereal is selected from at least one of the group consisting of: rice, oats, quinoa, wheat, and barley.

[0097] In one implementation, the nut is selected from at least one of walnuts, almonds, and pistachios.

[0098] In one implementation, the algae is selected from at least one of seaweed and microalgae.

[0099] In one implementation, the oilseed crop and / or its cake is selected from soybeans, rapeseed, peanuts, and bamboo.

[0100] In one implementation, the animal protein is selected from at least one of the following: milk, eggs, seafood, poultry, red meat, and white meat.

[0101] Suitable protein sources can be selected from any suitable animal or plant protein sources.

[0102] In one embodiment, the composition comprises a prebiotic source, which is at least one oligosaccharide.

[0103] Oligosaccharides can be isolated from any source. Preferably, oligosaccharides are isolated, purified, or concentrated from protein sources. Alternatively, all or some of the oligosaccharides are produced entirely or partially through bioengineering.

[0104] In one embodiment, the composition comprises at least one oligosaccharide selected from the group consisting of: fructooligosaccharides, inulin, guar gum, galactooligosaccharides, β-glucan, resistant starch, arabinoxylan, pectin, raffinose, polydextrose, agar, or combinations thereof.

[0105] In one embodiment, the composition comprises an effective amount of at least one protein source and at least one prebiotic source.

[0106] In one embodiment, the composition comprises a protein source administered to the subject in an amount of at least about 1 mg to at least about 2400 mg per kg of the subject's body weight.

[0107] In one embodiment, the composition comprises a prebiotic source administered to the subject in an amount of at least about 10 mg to at least about 1000 mg per kg of subject body weight.

[0108] In one embodiment, the composition comprises at least one protein source (which is fava bean) and at least one prebiotic source (which is fructooligosaccharide).

[0109] In one embodiment, the composition comprises:

[0110] - A minimum of about 1 mg to a minimum of about 2400 mg of fava bean protein per kg of the subject's body weight.

[0111] - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

[0112] In one embodiment, the composition comprises at least one protein source (which is pea) and at least one prebiotic source (which is fructooligosaccharide).

[0113] In one embodiment, the composition comprises at least one protein source (which is legume) and at least one prebiotic source (which is fructooligosaccharide).

[0114] In one embodiment, the composition comprises:

[0115] - A minimum of about 1 mg to a minimum of about 2400 mg of legume protein per kg of subject body weight.

[0116] - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

[0117] In one embodiment, the composition is used to prevent and / or treat age-related disorders.

[0118] In one implementation, the age-related impairment is selected from at least one of the following groups: physiological states or impairments related to cellular senescence, physiological states related to metabolic fatigue in one or more cells, excessive catabolism, oxidative stress, reduced antioxidant capacity and / or mitochondrial function, impaired activity, healthy period and / or lifespan of the subject, muscle dysfunction, including sarcopenia, weakness, inclusion body myositis, myopathy / rhabdomyolysis induced by drugs such as corticosteroids or statins, muscle wasting induced by inactivity or hospitalization, myasthenia gravis, immune dysfunction, cognitive impairment, including delirium, dementia, learning disabilities, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD), cardiovascular disease, impaired intestinal barrier integrity or leaky gut, impaired gut microbiome, and age-related inflammation.

[0119] In one embodiment, the composition is used to: (i) increase resistance to age-related symptoms in a subject; (ii) improve physiological states or disorders associated with cellular senescence; (iii) improve physiological states associated with metabolic fatigue in one or more cells; (iv) increase mitochondrial energy in one or more cells; (v) increase antioxidant capacity, reduce oxidative stress, and / or enhance mitochondrial function; (vi) improve activity and / or (vii) improve the subject's health and / or lifespan.

[0120] In one embodiment, the composition is in the form of a food composition, dietary supplement, nutritional composition, nutritional product, powdered nutritional product reconstituted in water or milk before consumption, food additive, medicine, beverage, and drink, including coffee-based products, oral nutritional supplements (ONS), complete nutritional formulas, pharmaceuticals, medical or food products.

[0121] In one implementation scheme, the subjects are humans, livestock, or pets.

[0122] Another aspect of this disclosure is the use of the composition in subjects who require at least one of the following: (i) increased resistance to age-related conditions; (ii) improved physiological states or disorders associated with cellular senescence; (iii) improved physiological states associated with metabolic fatigue in one or more cells; (iv) increased mitochondrial energy in one or more cells; (v) increased antioxidant capacity, reduced oxidative stress, and / or enhanced mitochondrial function; (vi) improved activity and / or (vii) improved individual health and / or lifespan.

[0123] The advantage of one or more embodiments provided in this disclosure is that they improve the condition of subjects, animals, humans, aged animals, or aged humans.

[0124] Another advantage of one or more embodiments provided in this disclosure is that it reduces or prevents the risk of morbidity or mortality due to excessive catabolism.

[0125] Another advantage of one or more embodiments provided in this disclosure is that it protects elderly subjects from muscle dysfunction, such as sarcopenia, weakness, inclusion body myositis, myopathy / rhabdomyolysis induced by drugs such as corticosteroids or statins, and muscle wasting induced by inactivity or hospitalization.

[0126] Another advantage of one or more embodiments provided in this disclosure is that it protects elderly subjects from the effects of myasthenia gravis.

[0127] Another advantage of one or more embodiments provided in this disclosure is that it protects aging subjects from cardiovascular disease or conditions.

[0128] Another advantage of one or more embodiments provided in this disclosure is that it protects aging subjects from impaired intestinal barrier integrity or leaky gut, damaged gut microbiome, and age-related inflammation.

[0129] Another advantage of one or more embodiments provided in this disclosure is that it improves the survival rate of critically ill patients or elderly subjects.

[0130] Another advantage of one or more embodiments provided in this disclosure is that it accelerates the recovery of activity or shortens the period of inactivity after leaving the intensive care unit.

[0131] The combination of the ingredients of this invention also effectively improves health. In addition, increased life expectancy is an indicator of improved health, as shown in Martineau CN, Brown AEX, Laurent P (2020) PLoS Comput Biol 16(7).

[0132] The following detailed reference is made to specific embodiments of the invention. While the invention will be described in conjunction with these specific embodiments, it should be understood that it is not intended to limit the invention to such specific embodiments. Rather, it is intended to cover alternatives, modifications, and equivalents that may be included within the spirit and scope of the invention as defined by the claims. Numerous specific details are set forth in the specification to provide a thorough understanding of the invention. The invention may be practiced without some or all of these specific details. In other instances, well-known methods and protocols have not been described in detail so as not to unnecessarily obscure the invention.

[0133] Composition formulation

[0134] In one embodiment, the composition is a food composition, including human food compositions and pet food compositions.

[0135] For pet food compositions, they provide the necessary dietary needs for an animal, an animal reward (e.g., biscuits), or a dietary supplement. The composition may be a dry composition (e.g., coarsely ground food), a semi-moist composition, a wet composition, or any mixture thereof. In another embodiment, the composition is a dietary supplement, such as gravy, drinking water, beverage, yogurt, powder, granules, paste, suspension, chewables, small pieces, rewards, snacks, pellets, pills, capsules, tablets, or any other suitable form of delivery. The dietary supplement is applied in small amounts, or in alternative embodiments, it may be diluted prior to application. The dietary supplement may need to be mixed, or may be mixed with water or other diluents prior to application.

[0136] In another embodiment, the composition is in the form of a food supplement or dietary supplement, particularly in the form of tablets, powder sachets, or gummies.

[0137] In a preferred embodiment, the composition is in the form of an effervescent tablet.

[0138] Effervescent tablets can be manufactured and controlled in the same way as conventional tablets. These controls include physicochemical properties such as hardness, weight variation, brittleness, dissolution time, pH, and content uniformity. Effervescent tablets can be prepared by direct compression, melt granulation, wet granulation, dry granulation, or any other suitable method. Low relative humidity in the environment (e.g., a maximum of 25% or less) and moderate to cold temperatures (e.g., about 25°C or 77°F) may be necessary to prevent granules or tablets from adhering to the tableting machine.

[0139] In direct compression, effervescent tablets can be formed by compressing powdered ingredients into dense blocks, for example, using a tablet press. Before tableting, the powdered ingredients can first be granulated to similar or equal sizes, resulting in a powder mixture with excellent flowability and no particle segregation. If the raw materials are selected to obtain a free-flowing, non-segregating, compressible powder mixture, granulation may not be necessary. The tablets can then be dried at a suitable temperature for a suitable time by heating, such as in an oven with air circulation, and after cooling, can be packaged in suitable packaging.

[0140] In the melt method, the ingredients can be mixed for an appropriate time in a suitable mixer, such as a co-mixer. The resulting mixture can then be heated to a suitable temperature. The powder can be mixed periodically until the water of crystallization of citric acid is released as a binder (e.g., about 30 minutes) and a suitable paste is obtained. This wet material can be sieved to obtain the desired granules, which can then be dried at a suitable temperature for an appropriate time. After drying, the granules can be sieved again. Other ingredients can be added to the granular material and mixed for an appropriate time. The granular mixture can then be compressed into tablets using a tableting machine. Finally, the tablets can be dried and packaged in suitable packaging.

[0141] In wet granulation, the ingredients can be ground individually or as a mixture with ethanol, an ethanol-water mixture, isopropanol, etc., using a mill, and the resulting powder can be sieved and then mixed. A binder solution can be added to the mixture to form a paste. This paste can then be sieved to obtain the desired granules and then dried. The dried material can be sieved again, and other ingredients can be added and mixed. The resulting granule mixture can then be compressed into tablets using a tableting machine. Finally, the tablets can be dried and packaged in suitable packaging. Wet granulation can also be carried out by carefully adding 0.1% to 1.0% (by weight) of water to a blend of raw materials that have the homogeneity, compressibility, and flowability required for producing high-quality tablets, but lack the required binding properties. Free water, typically added in the form of a fine spray to the selected formulation components while being mixed in a suitable mixer, acts as a binder. The granulation step must be precisely timed, and the ingredients must be thoroughly mixed to ensure that the granulation fluid is uniformly distributed in the blend. The mixture is then rapidly discharged into a drying oven. After drying, the granules are graded and then undergo final mixing. The granules are then compressed into tablets using a tableting machine.

[0142] Dry granulation methods utilize specialized processing equipment called "roll compactors" or "chilsonators." These machines compress premixed powders under extreme pressure between two opposing rotating rollers. Depending on the rollers' construction, if they have grooved or etched surfaces, the feed material can be compacted into dense strips of material called flakes (smooth rollers) or dense lumps (almond-shaped or rod-shaped). The compressed material is then reduced to a suitable size for tablet granulation purposes. Another dry granulation method is slugging, where powder particles are compressed into large, flat tablets or granules using a tablet compactor or, more commonly, heavy-duty tableting equipment. The resulting tablets or lumps are then ground to produce the desired granular characteristics.

[0143] Effervescent tablets can be made in any shape and can have any suitable size. As a non-limiting example, if they are round, the length, thickness, and / or diameter of the tablet can be 5 mm to 20 mm. The size of the tablet can be 5 mm to 10 mm, 5 mm to 15 mm, 10 mm to 15 mm, 10 mm to 20 mm, or 15 mm to 20 mm.

[0144] Effervescent tablets may contain a binder, such as polyvinylpyrrolidone (PVP) or any other suitable binder. The binder is preferably water-soluble. It can be added in dry powder or wet form as a aqueous or hydroalcoholic solution. Small amounts of mannitol, PEG 6000, and water can also be used as a binder. A 3% level of PEG 6000 can be used as a dry binder. The ideal amount of binder is one that makes the tablet hard enough to handle but soft enough to disintegrate and dry enough to be stable. Effervescent tablets can also be formulated without a binder.

[0145] Beverage composition

[0146] In one embodiment, the composition is a beverage composition. Such beverage compositions are intended for human or animal consumption. In several embodiments, the beverage is a dairy-containing beverage; a performance nutrition product; a medical nutrition product; a dairy product, such as a dairy beverage, containing at least one nutrient for improving motivational performance or mental energy.

[0147] dairy products

[0148] In one embodiment, the composition may be formulated with milk proteins as a “dairy product,” such as a milk protein concentrate or milk protein isolate; caseinate or casein, such as a micellar casein concentrate or micellar casein isolate; or whey protein, such as a whey protein concentrate or whey protein isolate.

[0149] Nutritional supplements

[0150] In one embodiment, the compositions of the present invention can be formulated as a "nutritional supplement" together with the glutathione-enhancing compounds of the present invention. The compounds of the present invention can be used alone or in combination with suitable additives to prepare tablets, gummies, powders, granules, or capsules, for example, in combination with conventional additives such as lactose, mannitol, corn starch, or potato starch. Binders, such as crystalline cellulose, cellulose functional derivatives, gum arabic, corn starch, or gelatin; disintegrants, such as corn starch, potato starch, or sodium carboxymethyl cellulose; lubricants, such as talc or magnesium stearate; and, if necessary, diluents, buffers, wetting agents, preservatives, and flavoring agents.

[0151] application

[0152] The compositions of the present invention may be administered at least one day per week, preferably at least two days per week, more preferably at least three or four days per week (e.g., every other day), and most preferably at least five, six, or seven days per week. The duration of administration may be at least one week, preferably at least one month, more preferably at least two months, and most preferably at least three months, for example, at least four months. In one embodiment, dosing is performed at least daily; for example, the subject may receive a dose once or more daily. In some embodiments, administration continues for the remainder of the individual's life. In other embodiments, administration occurs until detectable symptoms of the disease are retained. In a specific embodiment, administration continues until at least one symptom shows detectable improvement, and in other cases, administration continues to maintain remission.

[0153] The ideal duration of application of the composition can be determined by those skilled in the art.

[0154] Example

[0155] Example 1

[0156] A short-term, single-stage colonic simulation using ProDigest was used to study the effects of a protein and fiber mixture on the gut microbiota. Influence

[0157] Methods and Materials :

[0158] In vitro digestion :

[0159] The in vitro digestion simulation model followed the static digestion protocol of the international INFOKES consortium. In short, eight test products (Table 1) were administered entirely via oral, gastric, and small intestinal stages, the latter involving absorption via dialysis. Pre-digestion was considered important because all products contained a primary fraction of indigestible material that could be fermented by gut bacteria. ProDigest further refined this digestion method by combining more precise pH profiles with simulations of small intestinal absorption via dialysis. The latter enabled the removal of small molecules from the intestinal digestate using a 14 kDa dialysis membrane.

[0160] In vitro batch fermentation :

[0161] Fecal material was collected from five healthy adult donors within the framework of previous projects, but in this study only samples from three donors (A, B, and E) were used. Briefly, fecal suspensions were prepared and mixed with an internally optimized cryoprotectant. The resulting suspensions were aliquoted, rapidly frozen, and then stored at -80°C (cryostock).

[0162] Short-term screening assays involved colonic incubation of a single dose of the test compound using a bacterial inoculum from the donor under representative proximal colonic conditions. At the start of the short-term colonic incubation, 210 mg of FOS was added to the appropriate reactor (Table 2). Because the nutrients in sugar-depleted nutrient media would also be fermented by the colonic microbiota, a blank condition containing only sugar-depleted nutrient media (without FOS or pre-digested protein solution) was included for each donor. Fecal inoculum from donors A, B, and E was selected by the customer. All treatment groups and blanks are shown in Table 1.

[0163] Table 1. Treatment Groups

[0164]

[0165] Adding the prebiotic FOS to proteins increases spermidine production in the human gut microbiota.

[0166] Figure 1 (A) shows the results for spermidine after 24 hours of fermentation. The amount of spermidine was highly variable within each treatment, and some treatments (P5 and P7) resulted in very low amounts of spermidine, close to the blank control. Although higher values ​​than the blank control were observed in some groups (P1, P2, P3, P4, P6, and P8), there was no statistically significant difference in spermidine concentration when the protein digests were tested individually. Conversely, the addition of prebiotic FOS had a significant effect on spermidine production in the human gut microbiota, and algal proteins (P8) resulted in a significant increase in spermidine relative to the blank control, such as... Figure 1 As shown in (B).

[0167] FOS has a greater effect on spermidine (A) than on putrescine (B) produced by the gut microbiota. write .

[0168] Next, the inventors investigated whether the effect of FOS was spermidine-specific or other polyamine-specific. For example... Figure 2 As shown in (A), all proteomes exhibited higher levels of spermidine when FOS was added to the fermentation medium. Significant differences were found in P5 and P6, and an increasing trend was observed in the blank control, P2, and P7.

[0169] Unlike spermidine, under all conditions, the concentration of putrescine is at least one magnitude higher than that of spermidine, such as... Figure 2 As shown in (B). Moreover, compared with spermidine, protein digests and FOS have different effects on putrescine levels ( Figure 2 Even without FOS, all groups had detectable levels of putrescine. When FOS was added, the putrescine concentration remained unchanged in all groups.

[0170] When fermentation was extended to 24 and 48 hours, FOS rescued the positive relationship between arginine and spermidine in the protein. relation .

[0171] Arginine is a substrate for microbial polyamine production. The inventors next examined whether the amount of arginine in the protein source determines the polyamine concentration. Figure 3 As shown in Figure 5, the amount of arginine found in the protein source was positively correlated with the amount of spermidine after 6 hours of fermentation (R² = 0.72, p < 0.0001). Figure 3 (A)), and adding FOS does not change the relation (R2=0.44, p=0.0004, Figure 3 (A)). However, when fermentation was extended to 24 hours and 48 hours, at 24 hours (R2=0.1, p=0.14, Figure 3 (B) or 48h (R2=0, p=0.99, Figure 3 No relationship was found between arginine and spermidine in the protein in (C). Interestingly, FOS salvaged the positive relationship at both 24h and 48h. Figure 3 (B) and Figure 3 (C) indicates the importance of prebiotics in supplying the gut ecosystem for spermidine production.

[0172] Example 2

[0173] Comparison of plant and animal-derived proteins for polyamine production using short-term in vitro fermentation systems

[0174] Methods and Materials

[0175] In vitro digestion

[0176] Protein samples and negative control samples were first digested before being fermented by the human gut microbiota. The names of the protein samples from plants and animals are listed in Table 2. Notably, organic pea proteins from two different sources (Source 1: Hillpharma incorporate; Source 2: Puris) were tested and compared. Briefly, a three-step protocol simulated the digestive process in the mouth, stomach, and small intestine. In the oral administration phase, the test product was diluted in a simulated saliva liquid (1:1 wt / wt) and normalized to a final concentration of 0.04 g protein per mL. CaCl2 (1.5 mM) and amylase (75 U / mL) were added to the mixture, and the mixture was incubated at 37°C at pH 7.0 with stirring for 2 min. Because the protein products had different protein content ratios, adjustments were made to the weight of the protein material to normalize the protein load to 0.04 g / mL during the oral administration phase, except for the fava bean protein concentrate. The actual protein content in the broad bean protein concentrate was close to 60%, but the setting was adjusted to 80%, so the amount of protein tested was lower than ideal. Then, oral tablets were mixed with simulated gastric fluid in an equal volume (1:1). During the gastric phase, pepsin (2000 U / mL) and CaCl2 (0.15 mM) were added to the mixture, and then incubated at 37°C and pH 3.0 for 2 h. At the end of gastric digestion, the gastric chyme was mixed with simulated intestinal fluid at a 1:1 ratio. Bile salts (10 mM), CaCl2 (0.6 mM), and trypsin (trypsin activity 100 U / mL) were included in the small intestinal digested chyme. Digestion conditions were 37°C and pH 7.0 for 2 h.

[0177] Removal of free amino acids and small peptides and characteristics of the remaining material

[0178] Small peptides and free amino acids were removed by methanol precipitation. Samples obtained from in vitro digestion (IVD) were precipitated with 80% MeOH (at -20°C for 1 h) and separated into supernatant (S) and pellets (P) by centrifugation (13,000 × g, at 4°C for 15 min). The undigested (P) fraction was lyophilized and stored at -20°C prior to fermentation experiments.

[0179] Table 2. Protein samples used in in vitro digestion and fermentation experiments

[0180]

[0181] Fecal sample collection

[0182] Stool samples from healthy volunteers were collected according to a protocol approved by the Lausanne Ethics Committee (CER-VD) (Authorization No.: 2020-00304). Inclusion criteria were healthy participants aged 18 to 60 years who provided informed consent and were willing to adhere to clinical research protocols. Exclusion criteria were: 1) adherence to specific dietary protocols, such as strict vegetarianism, veganism, ketogenic diets, or ancient diets; 2) chronic or recurrent diarrhea with spontaneous bowel movements more than twice a day; 3) antibacterial / antifungal treatment during the 3 months prior to study recruitment; 4) medications or supplements known to alter gut function or gut microbiota during the 4 weeks prior to study recruitment (i.e., anti-acid secretion agents, prebiotic / probiotic supplements, laxatives); 5) prior gastrointestinal surgery; 6) alcohol intake exceeding 2 servings per day; 7) intake of artificially sweetened beverages exceeding 1000 mL / day; and 8) current or historical gastrointestinal disease.

[0183] The preparation of fecal samples for in vitro fermentation followed the method described by Van den Abbele et al. In short, freshly collected fecal samples were placed in an airtight container equipped with AnaeroGen™ to minimize exposure to ambient oxygen. Once in the anaerobic chamber (Coy Laboratory Products, Grass Lake, MI, USA), the fecal material was diluted 10-fold (w / v) in anaerobic phosphate buffer containing 10% glycerol (0.1 M NaH₂PO₄ and 0.1 M Na₂HPO₄, in a 2:1 ratio), and aliquots (25 mL) of the undiluted fecal solution were stored at -80°C for later use.

[0184] In vitro fermentation

[0185] To prepare for the batch fermentation experiments, frozen fecal samples were thawed in a 37°C water bath for 5 minutes. The samples were then centrifuged at 2000 x g for 2 minutes at room temperature, and 250 μL of the supernatant was mixed with 10 mL of fermentation medium in sterile hungate tubes (composition shown in Table 3). In the hungate tubes, MeOH-precipitated pellets were resuspended in the fermentation medium at a final concentration of 3 mg / mL. Anaerobically fermentation was carried out at 37°C for 48 h during screening and 24 h in the second round of experiments. Each protein component was tested in three independent fecal samples to allow for statistical analysis.

[0186] Table 3. Composition of fermentation medium

[0187]

[0188] Quantitative analysis of polyamines in fermentation media

[0189] Quantifications of guanidine and other polyamines are documented in the internal analysis report (R0000880953). All samples were prepared according to internally developed analytical methods reported in R&D Memo: “Polyamines in bacterial media – Analytical approach using LC-HRMS”. For details on analytical methods and method validation, please refer to the above description and documentation.

[0190] In protein testing, broad bean protein produced the highest amount of spermidine, followed by pea protein, among which other proteins... It did not result in different amounts of spermidine compared to the control, and the protein test had no effect on the amount of putrescine. .

[0191] The concentrations of spermidine and putrescine at 24 h are shown in the figures below. Figure 4 (A) and Figure 4 (B) Of all the proteins tested, broad bean protein resulted in the highest amount of spermidine, followed by pea 870, while other proteins did not result in a different amount of spermidine compared to the control (enzyme only). Figure 4 (A)). In contrast to spermidine, the amount of putrescine was similar in all groups ( Figure 4 (B)).

[0192] Starch and fiber are expected to be found in the non-protein fraction. As the inventors observed in Example 1, FOS enhances spermidine production, and it is reasonable to expect that starch or fiber in broad beans will also promote spermidine synthesis in the same way as FOS.

[0193] The inventors have discovered that fiber (FOS) and a less refined protein source—fava bean protein concentrate—are promising nutritional candidates for enhancing the production of the anti-aging molecule spermidine.

[0194] The inventors also anticipate that other protein-prebiotic combinations or less refined protein formulations derived from soybeans, peas, lentils, or other plant sources will have similar advantages over animal proteins used for spermidine production.

[0195] Studies have shown that the intake of a combination of protein and prebiotic sources increases spermidine production in the gut microbiota.

[0196] It should be understood that various changes and modifications to the currently preferred embodiments described herein will be apparent to those skilled in the art. These changes and modifications can be made without departing from the spirit and scope of the subject matter of the invention and without diminishing its intended advantages. Therefore, such changes and modifications are intended to be covered by the appended claims.

Claims

1. A composition comprising at least one protein source and at least one prebiotic source, said composition being used to increase spermidine production in the gut microbiota of a subject.

2. The composition according to claim 1, wherein the protein source is at least one selected from the group consisting of: legumes, grains, nuts, algae, oil crops and / or their cakes, animal proteins or combinations thereof.

3. The composition according to claim 2, wherein the legume is selected from at least one of the group consisting of: broad beans, chickpeas, green beans, and lentils.

4. The composition according to claim 2, wherein the cereal is selected from at least one of the group consisting of: rice, oats, quinoa, wheat, and barley.

5. The composition according to claim 2, wherein the nut is selected from at least one of walnuts, almonds, and pistachios.

6. The composition according to claim 2, wherein the algae is selected from at least one of seaweed and microalgae.

7. The composition according to claim 2, wherein the oilseed crop and / or its pressed cake is selected from soybean, rapeseed, peanut, and bamboo.

8. The composition according to claim 2, wherein the animal protein is selected from at least one of the following: milk, eggs, seafood, poultry, red meat, and white meat.

9. The composition according to any one of the preceding claims, wherein the prebiotic source is at least one oligosaccharide.

10. The composition according to claim 9, wherein the at least one oligosaccharide is selected from the group consisting of: fructooligosaccharides, inulin, guar gum, galactooligosaccharides, β-glucan, resistant starch, arabinoxylan, pectin, raffinose, polydextrose, agar, alginate, or combinations thereof.

11. The composition according to any one of the preceding claims, wherein the composition comprises an effective amount of the at least one protein source and the at least one prebiotic source.

12. The composition according to any one of the preceding claims, wherein the protein source is administered to the subject in an amount of at least about 1 mg to at least about 2400 mg per kg of the subject's body weight.

13. The composition according to any one of the preceding claims, wherein the prebiotic source is administered to the subject in an amount of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

14. The composition according to any one of the preceding claims, wherein the at least one protein source is broad bean and the at least one prebiotic source is fructooligosaccharide.

15. The composition according to claims 8 and 14, wherein the composition comprises: - A minimum of about 1 mg to a minimum of about 2400 mg of fava bean protein per kg of the subject's body weight. - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

16. The composition according to any one of claims 1 to 13, wherein the at least one protein source is legumes, and the at least one prebiotic source is fructooligosaccharides.

17. The composition of claim 10, wherein the composition comprises: - A minimum of about 1 mg to a minimum of about 2400 mg of legume protein per kg of the subject's body weight. - A quantity of fructooligosaccharides of at least about 10 mg to at least about 1000 mg per kg of the subject's body weight.

18. The composition according to any one of the preceding claims, wherein the composition is used for the prevention and / or treatment of age-related disorders.

19. The composition of claim 18, wherein the age-related disorder is selected from at least one of the following groups: physiological states or disorders related to cellular senescence, physiological states related to metabolic fatigue in one or more cells, excessive catabolic metabolism, oxidative stress, reduced antioxidant capacity and / or mitochondrial function, impaired activity, healthy period and / or lifespan of the subject, muscle dysfunction, including sarcopenia, weakness, inclusion body myositis, myopathy / rhabdomyolysis induced by drugs such as corticosteroids or statins, muscle wasting induced by inactivity or hospitalization, myasthenia gravis, immune dysfunction, cognitive dysfunction, including delirium, dementia, learning disabilities, attention deficit disorder (ADD) and attention deficit hyperactivity disorder (ADHD), cardiovascular disease, impaired intestinal barrier integrity or leaky gut, impaired gut microbiome, and age-related inflammation.

20. The composition according to any one of the preceding claims, wherein the composition is used for: (i) increasing resistance to age-related symptoms; (ii) improving physiological states or disorders associated with cellular senescence; (iii) improving physiological states associated with metabolic fatigue in one or more cells; (iv) increasing mitochondrial energy in one or more cells; (v) increasing antioxidant capacity, reducing oxidative stress, and / or enhancing mitochondrial function; (vi) improving activity and / or (vii) improving the health period and / or lifespan of a subject.

21. The composition according to any one of the preceding claims, wherein the composition is in the form of a food composition, dietary supplement, nutritional composition, nutritional product, powdered nutritional product reconstituted in water or milk before consumption, food additive, medicine, beverage, and drink, including coffee-based products, oral nutritional supplements (ONS), complete nutritional formulas, pharmaceuticals, medical or food products.

22. The composition according to any one of the preceding claims, wherein the subject is a human, livestock, or pet.

23. Use of the composition according to any one of the preceding claims in a subject requiring at least one of the following: (i) increased resistance to age-related conditions; (ii) improved physiological states or disorders associated with cellular senescence; (iii) improved physiological states associated with metabolic fatigue in one or more cells; (iv) increased mitochondrial energy in one or more cells; (v) increased antioxidant capacity, reduced oxidative stress, and / or enhanced mitochondrial function; (vi) improved activity and / or (vii) improved individual health and / or lifespan.

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