Compositions and methods for resetting biology of cells

Abstract

A composition for administering to a subject includes a first compound chosen from S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B12, glycine, serine, threonine, and any combination thereof, and a second compound chosen from spermidine, spermine, calcium peroxide, zinc, N-Acetylcysteine, cysteine, α-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, O3, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaSH), sodium sulfide (Na2S), metformin, acetaminophen, curcumin, quercetin, ginseng, R-alpha-lipoic acid, apigenin, fisetin, melatonin, ginger, astaxanthin methylene blue and optionally, a carrier, and any combination thereof

Description

COMPOSITIONS AND METHODS FOR RESETTING BIOLOGY OF CELLSCROSS REFERENCE TO PRIOR APPLICATION

[0001] This application claims the benefit of U.S. Provisional Patent Application No.63 / 563,987 entitled as “Compositions and Methods For Resetting Biology of Cells”, filed March 12, 2024, which is incorporated by reference in its entirety.FIELD OF THE INVENTION

[0002] This disclosure generally relates to compounds and methods for reversing epigenetic aging by inducing changes within the Metabalome of human cells.BACKGROUND

[0003] Not all genes are expressed in all cells. This “selective gene expression” control of the 21,800 genes that code for proteins in human cells determines if the cell becomes a brain cell or a heart cell. This system of gene regulation is referred to as “epigenetics” (Kundaje A 2015). Epigenetics controls the rate of aging (Reynolds L 2014). One of the methods by which genes are regulated is the methylation of certain DNA residues called “cytosines.” In 1967, Boris Vanyushin showed that DNA loses its methylation with aging (Vanyushin B F 2005). There are other epigenetic mechanisms involved besides DNA methylation, such as historic protein modifications, microRNA, and chromatin remodeling (heterochromatin vs euchromatin) (Kundaje A 2015). In addition, some DNA cytosines increase their methylation with aging and other sites decrease their methylation with aging. It is clear that DNA methylation is the form of epigenetic gene regulation that correlates with aging. This has most recently been shown by Steven Horvath, who showed that a “DNA methylation clock” can be constructed from the analysis of only 353 cytosine residues and that this “DNAm clock” (Bocklandt S 2014) has a 0.96 correlation with aging. More importantly, this “clock” keeps time much better than any other known measure of aging besides birth date. DNA methylation profiling of mesenchymal stem cells (MSCs) obtained from individuals aged from 2 to 92 years identified 18,735 hypermethylated and 45,407 hypomethylated CpG sites associated with aging. Most importantly, hypom ethylated CpG sites were strongly enriched in the active chromatin mark H3K4mel in stem and differentiated cells, suggesting this is a cell type-independent chromatin signature ofDNA hypomethylation during aging. These results indicated that the dynamics of DNA methylation during aging depends on a complex mixture of factors that include the DNA sequence, cell type, and chromatin context involved and that, depending on the locus, the changes can be modulated by genetic and / or external factors (Fernandez A F 2015). It has been shown that calorie restriction prevents the age-related changes in DNA methylation in mice (Chouliaras L 2012). Two of the 7 Sirtuin enzymes have been shown to indirectly affect DNA methylation through their effects on histone deacetylation (Sirtl and Sirt6). It was also shown that the end-product of the Sirtuin reaction, nicotinamide, needs to be methylated to 1- methylnicotinamide, otherwise the end product nicotinamide will bind inside the Sirtuin enzyme and stop its enzymatic activity (Schmeisser K 2013). Sirtuin-1 decreases the activity of NF-kB which increases tri-methylation of lysine 36 on histone 3 (H3K36me3). This correlates to accelerated DNA methylation. Genes with a dramatic expression change during aging are marked with low or even undetectable levels of H3K36me3 in their gene bodies irrespective of their corresponding mRNA abundance (Pu M 2015). In human cells a global loss of trimethylation of H3K9 (H3K9me3) recapitulates accelerated cellular senescence and changes in heterochromatin architecture. These findings also correlated to people ages 7 to 72 heterochromatin's disorganization with increasing age (Zhang W 2015). In Jan. 30, 2015 DNA- methylati on-age of blood was used to predict all-cause mortality in later age of humans independently of health status, lifestyle factors and known genetic factors (Marioni R E 2015). On Feb. 19, 2015 the journal Nature published the results of 111 human epigenomes allowing future comparisons and references to be made by others (Kundaje A 2015).

[0004] Methionine is particularly sensitive to oxidative modification. Methionine is the step after homocysteine synthesis and prior to SAM synthesis in the methylation pathway. Thus methionine is a part of the methylation pathway and is regulated by redox balance. Cysteine is synthesized from methionine and is the main precursor of hydrogen sulfide (H2S). Elevated homocysteine levels are associated with inhibition of endogenous hydrogen sulfide (H2S) generation (Tang X 2011). Hydrogen sulfide (H2S) ameliorates methionine induced oxidative stress (Tyagi N 2009). Homocysteine (Hey) can be irreversibly degraded to hydrogen sulfide (H2S) by a transsulfuration pathway which is activated by oxidative stress. H2S has protective functions in hyperhomocysteinemia (Ohashi, R. 2005, Chang L 2008). Adipose tissue is an important organ of methionine metabolism and is also an insulin-sensitive organ. Increases ofH2S in adipose tissue increases insulin sensitivity (Feng X 2009). High pancreatic H2S suppresses insulin release (Wu L 2009). Blood levels of H2S are lower in type 2 diabetes than age matched healthy subjects (Jain S 2010). Aspirin is an arachidonate inhibitor and may influence the methionine-homocysteine cycle and associated one carbon metabolism and thereby both methylation and redox balance (Lupoli R 2015). H2S therapy with H2S donors; Na2S or NaSH inhibits aspirin in a dose dependent manner (Zanardo R C 2006).

[0005] There is also a methylation inhibitor: S-adenosylhomocysteine (SAH). When methionine is abundant, NNMT regulates only SAH not SAM (Ulanovskaya O A 2013).

[0006] As a general rule age-related hypo-methylation of DNA is the dominant event leading to increased expression of genes, but hyper-methylation is common in some promoter regions of DNA with age leading to promoter repression. There is a close relationship between redox balance and methylation balance (Metes-Kosik N 2012).

[0007] There is a relationship of methylation to redox balance with homocysteine going to the antioxidant glutathione when under oxidative conditions and going to SAM and methylation under reduction conditions (Mosharov E 2000).

[0008] Changing the redox potential of oxidation-sensitive protein thiols can allow the switch between distinct catabolic and anabolic processes as well as activate survival pathways. Protein methionine and cysteine residues are particularly sensitive to oxidative modification. Methionine is the step prior to SAM synthesis in the methylation pathway. Thus methionine is connected to the methylation pathway and regulated by redox balance. The percentage of cysteine residues increases with organism complexity but their prevalence are still significantly lower than occurrence simply based on codon usage. Cysteines that occur in clusters are highly conserved in evolution and usually are structurally or functionally important. pKa values for the thiol groups are influenced by their local environment. Oxidation states can range from the fully reduced thiol / thiolate anion to the fully oxidized sulfonic acid (Cremers C M 2013). The reaction rate of protein thiols with oxidants such as hydrogen peroxide (H2O2) spans over 7 orders of magnitude without any detectable correlation to the acidity of the respective active site thiol (Ferrer-Sueta G 2011).

[0009] There are reversible and irreversible cysteine modifications. Oxidation of cysteine thiol (RSH / RS) by ROS, RNS, or RCS leads to the formation of highly reactive sulfenic acid (RSOH), which can react either with another thiol to form a disulfide bond (RSSR) or with GSH to become S-glutathionylated (RSSG). These oxidative modifications are reversible, and reduction is catalyzed by the Trx and / or Grx system. Further oxidation of sulfenic acid to sulfinic acid (RSO2H) and sulfonic acid (RSO3H) is thought to be generally irreversible in vivo. Many of the thiol redox regulated proteins act as transcriptional regulators (e.g., OxyR, Yaplp) which rapidly induce expression of genes involved with antioxidant defenses (Zheng M 1998, Tachibana T2009), others are involved with signal transduction cascades (Gopalakrishna R 2000 and Dinkova-Kostova A T 2005). (See Supplement 1 from Cremes C M 2013 for more examples.)

[0010] An example of an enzyme with thiols in the active site is GAPDH which plays a crucial role in glycolysis. Oxidation of the GAPDH thiols blocks glycolysis and contributes to the generation of NADPH instead of NADH (Shenton D 2003). Another example is the oxidation of thiols in active sites that inactivate the phosphatase activity of SHP1 / 2, PTEN, Cdc25 enhancing signaling intensity achieved by substrate phosphorylation, this leads to activation of signaling pathways like NF -kB -inducing kinase / lKB, causing expression of genes involved in antioxidant defense (Jung K J 2009). A third example is the Sirtuin thiol groups in Sirtuin active sites that are very sensitive to oxidation, which inhibit Sirtuin activity when oxidized. Human Sirtuin- 1 has 3 (Cys-67, Cys-268, and Cys-623) of 5 cysteines exposed to possible reversible thiol modification via redox balance (Autiero I 2008). Cys-67 and Cys-623 are consistent with post- translational regulation of these terminal regions, Cys-268 lies in the NAD+ binding region in all the members of the Sirtuin family of which the catalytic core is highly conserved. The binding of NAD+ results in changes in the Sirtuin conformation that allows catalysis to proceed (Zee R2010).

[0011] There are different types of ROS and RNS (reactive nitrogen species). Together they are referred to as RONS. They include: superoxide, hydrogen peroxide, hydroxyl radicals, singlet oxygen, nitric oxide, peroxynitrite, hyperchlorite, and also lipid peroxidations “PUFA”s. There is different specificity of ROS. ROS display a type of specificity that is atomic rather than molecular. ROS most often reversibly reacts in cell signaling with Sulphur, which is one of the least abundant atoms in biological macromolecules and mostly with side chains of cysteine ormethionine residues in peptides or proteins (Nathan C 2013). Endogenous enzymatic sources of ROS (multiple isoforms allow more sensitivity and specificity in regulation) include seven isoforms of NADPH oxidases (NOXs) that are differentially expressed (regulated) in diverse cells and species as well as a list of other sources (see Box 1 page 2 Nathan C 2013).

[0012] The main types of anti-oxidants enzymes (multiple isoforms allow more sensitivity and specificity in regulation) (need control and use of transition metals) are Superoxide Dismutase (SOD) 3 isoforms, Glutathione Peroxidase (GPX) 5 isoforms, and Catalase. There are also Thioredoxin (TRX) 2 isoforms (with thioredoxin reductase) Thioredoxin may be recycled by interaction with REF-1 (REF-1 keeps Sirtuin thiols reduced), Glutaredoxin (GRX) 3 isoforms, Peroxiredoxin (PRX) 6 isoforms (responsible for reduction of 90% of Eukaryote mitochondrial and more than that of cytosol H2O2. This can be turned on and off with a functional loop of regulation allowing redox signaling (Sies H. 2014). Peroxiredoxin makes up a phylogenetically ancient family of proteins whose primary role is detoxification of H2O2. These also create a redox rhythm. It is thought that catalytic cycle of peroxiredoxin hyperoxidation and recycling by sulfiredoxin may form the basis of a transcription-independent circadian clock (Rey, G. 2013). NAD+ levels are correlated to the biological clock with 2 peaks in the day 12 hours apart. It is thought that because NAD+ / NADH with a cellular ratio of >1 is higher than NADP+ / NADPH cellular ratio of <0.01 in the cytosol that this allows the cell to segregate antioxidant and biosynthetic reducing equivalents (NADPH) from those destined for mitochondrial ATP generation (NADH). The phosphate of NADPH confers different substrate specificity but has the same electron transfer properties. Hyperoxidation of peroxiredoxins can induce chaperone function as well as signal transduction.

[0013] Antioxidant small molecules include: Glutathione (GSH), Uric acid, Bilirubin, Ascorbic Acid (Vitamin C) Vitamin E, also carotenoids, Co-Enzyme Q10, N-acetylcysteine (NAC). NAC acts as a reduced thiol donor and counters H2O2 that oxidizes thiols.

[0014] In 1935, Clive McCay first discovered that caloric restriction (CR) increased life span in animals. CR is the practice of reducing caloric consumption without inducing malnutrition. This requires an organism receive adequate amounts of water, vitamins, minerals, and protein, but limits carbohydrate and fat calories (to less than the recommended dietary allowance (RDA) forhumans). CR can be done safely without harmful health effects with total caloric restrictions in the range of 10-40% less than RDA recommendations. In 1986, Richard Weindruch showed that restricting calories to Ards of the normal amount in mice increased lifespan by 40%. To date, a large number of experiments in animal models have corroborated these results. Animal models of CR have also helped researchers discover the molecular biology pathways that account for the increase in life span and health span (Colman R J 2014). A randomized controlled two year calorie restriction study in humans (Ravussin E 2015) showed feasibility and effects on predictors of health-span and longevity (life-span).

[0015] Iron is also an essential element for living cells. However, as people age, the amount of free iron within the cell increases, as iron storage proteins such as ferritin can be damaged and leak free iron into the cell (Mangan D 2021). Superoxides may also damage ferritin, resulting in additional iron leakage. Furthermore, it has been found that blocking the absorption of iron in research animals such as Drosophilia extends their lifespan (Massie HR 1993). Other studies have found that life extending compounds interact with iron by either chelation, absorption inhibition, or general iron loss (Mangan D 2021).SUMMARY

[0016] Biological and cellular age is typically assessed using a number of biomarkers, including telomere length, cellular senescence biomarkers, and DNA methylation. Of these, DNA methylation is an ideal surrogate for epigenetic age.

[0017] In accordance with the purposes of the disclosed materials and methods as embodied and broadly described within, the disclosed subject matter disclosure generally relates to compounds and methods for reversing epigenetic aging by inducing changes within the Metabalome of human cells. Primarily through the use of a methyl donor such as methionine, and the source of an oxidant such as spermidine, which also donates an amino butyl group for the synthesis of hypusine. Threonine may also be included to synthesize additional methionine and up regulates ferritin to protect the cell from ferroptosis.

[0018] S-5'-adenosyl-L-methionine (SAM), betaine, choline, folate, vitamin B 12, glycine, serine, and threonine can be used in combination with, or in place of methionine. Spermine, calcium peroxide, zinc, N-Acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine,ergothioneine, urolithin A, terpenoids, berberine, 021, 02, H2S, 03, H0C1, HOBr, HOI, Na2S, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaHS), sodium sulfide (Na2S), diallyl trisulfide (DATS), metformin, acetaminophen, ortho hydroxyphenols, para dihydroxy phenols, R00H, ROS, RNS, RCS, RSOH, ROOM, where R is alkyl, cycloalkyl, heteralkyl, heterocycloalkyl, alkenyl, heteroalkenyl, cycloalkenyl, or hetercycloalkenyl, and other oxidants can be used in combination with, or in place of spermidine to promote and activate sirtuins and other enzymes that repair DNA.

[0019] The compound is typically administered over multiple hours and multiple days, typically in conjunction with fasting.

[0020] WO / 2017 / 062311 provides composition and methods which can be used for human age reversal therapy and WO / 2021 / 202245 provides compositions and methods for treating viral infections.BRIEF DESCRIPTION OF THE DRAWINGS

[0021] FIG. 1 is an illustration of a pathway connection showing the link between category 1 compounds and category 2 compounds.

[0022] FIG. 2 is a biological age test result on a subject.

[0023] FIG. 3 is an illustration of an administration timeline in an embodiment of the disclosed subject matter.DETAILED DESCRIPTION

[0024] The disclosed subject matter includes a composition and method of treatment intended to reduce aging, treat age-related diseases, or similar conditions. The composition includes two categories of compounds that complement one another in the subject. The first category, referred to herein as category 1, includes compounds involved in methylation or methylation pathways within the subject. An example of methylation is when a compound donates a methyl group to DNA, RNA, or proteins. Various compounds participating in methylation promote the synthesis of methionine, a precursor compound involved in methylation pathways, including methylationof DNA and RNA. Methylation influences numerous biological processes, including the regulation of gene expression.

[0025] The second category of compounds, referred to herein as category 2, includes compounds that control or reduce oxidative stress in a subject. Certain compounds control oxidative stress by directly targeting reactive oxygen species (ROS). Other compounds indirectly control oxidative stress by supporting pathways involved in the reduction of reactive oxygen species. Additionally, certain category 2 compounds reduce the generation of reactive oxygen species within the body. Numerous compounds may regulate oxidative stress by various pathways, and accordingly, many such compounds may fall within category 2 of the disclosed composition.

[0026] The disclosed method of treatment includes administering a composition comprising at least one category 1 compound and at least one category 2 compound. Multiple category 1 and / or category 2 compounds may also be combined in a single composition.

[0027] In various embodiments, the composition may be administered to the subject in pulses. A pulse is a dosing period during which the subject is administered doses of the composition containing at least one category 1 compound and at least one category 2 compound over a defined period of time. For example, a subject may be administered the composition four times per day for several consecutive days. In an embodiment, the subject may be administered doses periodically over four consecutive days. In various embodiments, the subject may fast prior to the dosing period. In embodiments, a pulse may include a fasting period followed by a dosing period, followed by a cessation period during which the subject does not fast and does not receive any doses of the composition. The number and frequency of pulses administered to a subject may vary. For example, a subject may receive one pulse per month. In another example, a subject may receive four pulses over the course of one year.

[0028] Disclosed herein extended pulse lengths of two categories of compounds when used together provide a variety of beneficial effects, such as biological clock age reversal, to mammals, including humans. An example of a category 1 compound is methionine, which is precursor to S-adenosyl-methionine, and an example of a category 2 compound is spermidine. In addition, to methionine and spermidine, other compounds can be added such as glycine orthreonine. Between 0.1 mmole / kg human body weight / day to 1 .0 mmole / kg human body weight / day of these components is optimal. It is optimal to dose 4 times per day every 5 hours, or every 4 hours, or every 6 hours. It is preferable to dose at least 0.1 mmole / kg human body weight, 0.2 mmole / kg human body weight 0.4 mmole / kg human body weight 0.6 mmole / kg human body weight, or 0.8 mmole / kg human body weight. It is preferable to dose not more than 0.1 mmole / kg human body weight, 0.2 mmole / kg human body weight 0.4 mmole / kg human body weight 0.6 mmole / kg human body weight, or 0.8 mmole / kg human body weight.

[0029] In one method, a composition of methionine and spermidine are provided to a subject in a pulse of at least 1,2, 3, 4, 5, or 6 days. Between 1 to 21 days is optimal for the total duration for the pulse. It is optimum to fast only with water during this pulse. Preferably at least 4 days, 5 days, 6 days, 7 days, 8 days, 10 days, or 15 days. Preferably not more than 4 days, 5 days, 6 days, 7 days, 8 days, 10 days, or 15 days.

[0030] For the oxidative pulse, the oxidants previously used (H2O2 and H2S) are shorter lived, so spermadine can provide a more stable category 2 compound. Taurine provides a slower release of H2S donors or can use cysteine instead, a- ketoglutarate can be used as well since it makes cysteine which in turn can make H2S. Spermidine makes H2O2 in the cell and Taurine makes H2S in the cell as does cysteine.

[0031] Because there is now a longer oxidative pulse, you can increase protection of the cell from cell suicide from the oxidation due to available iron which has become available (See page 6 on Schuler AA 2022 for list of papers) by adding threonine which can help protect against this (Kim I 2022).

[0032] Threonine up regulates (increases) the ferritin protein. With age and with senescence the iron storage capacity of ferritin is compromised. This means there is more iron escaping from ferritin, thus more iron available to increase ROS (reactive oxygen species). Less available iron in the cytoplasm correlates to a longer and healthier life (Kao Y-R 2024). This increase in ROS correlates to a decrease in glutathione, a small molecule anti-oxidant. Glutathione can be increased with N-acetylcysteine and glycine. Threonine promotes the inhibition of ferritindependent “ferroptosis” which is a protective form of regulated cell death similar to apoptosis. (James SA 2015, Jenkins NL 2020)

[0033] One can use “GRIM Age” DNA methylation clocks as the primary surrogate for a biological age marker and “Inflammaging” cytokines (such a IL-ip or IL-6) as the secondary surrogate markers in this human study to determine if human biological age reversal has happened. DNA methylation age clocks are the same for 59 different tissue types and 185 mammalian species (Lu AT 2023). Epigenetic aging, which is measured here with GRIM Age (version 2) is associated with nutrient sensing, mitochondrial activity and stem cell composition (Kabacik S 2022). The peak expression of genes in re-methylation reprogramming in the mouse happens between day 4 and 6 (from line 243), the mouse reprogramming process takes about 14 day while the human reprogramming process is similar (not identical) and takes longer, about 30 days (from line 334) (Kriukov D 2022) in BioRxiv December 14 2022 and (Fu K 2018). This reprogramming process needs and starts with transient genome-wide de-methylation (Pasque V 2011, Apostolou E 2013, Theunissen TW 2014) effecting H3K9me3 as well. De-methylation starts with the oxidation of 5-methyl Cytosine and the loss of H3K9me3, most likely due to erosion of methylation and lack of repair. The cell has a record of where the methylation should go and re-methylates on its own after the de-methylation step has taken place and ended (Sinclair D Lifespan book 2019 and Lu Y 2020). This happens, following the stopping of the “4 to 6 day Therapy” and benefits continually increase with improvements continually adding for approximately 5 weeks (2023 ARDD Re-methylation lecture in Copenhagen by Sharon Rosenzweig-Lipson PhD).

[0034] The disclosed subject matter includes a composition made of at least one category 1 compound and at least one category 2 compound. The category 1 compounds generally support and facilitate cellular methylation. The process of cellular methylation modifies DNA. In an example, a modification to DNA may result in regulating gene expression. Among other things, gene expression may influence the function of cells.

[0035] Examples of category 1 compounds include methionine as well as S- adenosylmethionine, or SAM. SAM may donate methyl groups and become S- adenosylhomocysteine. S-adenosylhomocysteine may convert to homocysteine and regenerate methionine. The regeneration of methionine may occur via vitamin B 12 and folate. Additional examples of category 1 compounds may include betaine and choline. Betaine may donate methyl groups to convert homocysteine to methionine. Choline may serve as a precursor to betaine.

[0036] The category 1 compounds generally act as methyl donors. Methyl donors in metabolic pathways may facilitate synthesis of intermediates in the metabolic pathways. Accordingly, the category 1 compounds act to fortify various metabolic pathways by synthesizing the intermediates that propagate through the various pathways. Likewise, the category 1 compounds may also regulate epigenetic markers.

[0037] Category 1 compounds may provide methyl groups that form epigenetic markers.Epigenetic markers are chemical modifications that influence gene expression. The category 1 compounds support maintenance such as DNA methylation. DNA methylation is the chemical marking of DNA that effectively regulates gene expression on the DNA. The category 1 compounds may further regulate histone methylation patterns. Histones form a chemical structure around which DNA wraps, also controlling gene expression. Accordingly, category 1 compounds, through DNA methylation and histone methylation, may influence gene silencing and gene activation. The category 1 compounds may also control chromatin structure and availability.

[0038] The combination of a category 1 compound and a category 2 compound in the composition may complement one another. The category 2 compounds, in general, may facilitate and modulate oxidative pathways. The combination of category 1 and category 2 compounds may balance methylation and oxidative signaling. The balancing of methylation and oxidative signaling may improve cellular function, cellular response, as well as gene signaling and gene expression.

[0039] Various amounts of a compound of category 1 may be used in combination with a compound of category 2 to effectuate treatment in a subject. The amounts and concentrations of category 1 and category 2 compounds in the disclosed composition are disclosed herein. For example, various amounts of category 1 and category 2 compounds are provided herein. In an embodiment, the composition may be administered one or more times per day for a number of days, such as three to four days, and then stopped for a number of days before starting again.

[0040] The composition of category 1 and category 2 compounds, when administered, may demonstrate measurable changes in various markers of the subject’s health. For example,administration of the composition in subject may reduce the biological age of cell samples taken from the subject. Administration of the composition may reduce inflammation in the subject.

[0041] Administration of the composition may demonstrate changes in DNA methylation patterns. For example, gene expression at various gene sites may be shown to be modified based on changes in DNA methylation. Various methods for determining or measuring DNA methylation include pyrosequencing, DNA methylation arrays, and targeted methylation-specific sequencing.

[0042] The disclosed subject matter includes a composition comprising at least one category 1 compound and at least one category 2 compound. The category 2 compounds generally facilitate oxidative processes in a subject. For example, the category 2 compounds may generate reactive species that produce reactive oxygen species in the subject. In another example, a category 2 compound may modify oxidative stress responses in a subject. In various examples, the category 2 compounds may indirectly reduce oxidative stress in a subject, such as by modulating cellular antioxidant pathways.

[0043] In embodiments, the category 2 compound may include spermidine. Spermidine may cause or control reactive oxygen species in a subject. Spermidine may act to reduce cellular oxidative stress by clearing damaged mitochondria and misfolded proteins. Spermidine may also reduce oxidative damage in cells by reducing oxidized components in the cell. In embodiments, the category 2 compound may include taurine. Taurine may control oxidative stress indirectly by producing hydrogen sulfide in a subject. Taurine may reduce inflammation in a cell. Taurine may further stabilize membranes in cells. Additionally, taurine can directly reduce oxidative stress. In embodiments, the category 2 compound may include cysteine. Cysteine may indirectly control oxidative stress by facilitating synthesis of hydrogen sulfide, which may affect signaling in a subject.

[0044] The category 2 compounds listed herein in the disclosed subject matter may facilitate oxidative pathways. For example, the category 2 compounds listed herein generally facilitate oxidative signaling in the subject. In another example, category 2 compounds may regulate oxidative stress responses by either reducing or generating reactive oxygen species. In some cases, generation of reactive oxygen species by category 2 compounds may assist in balancingoxidative stress in the subject. Additionally, category 2 compounds may maintain oxidative and redox balance at the cellular level. They may further prevent oxidative damage at the cellular level.

[0045] All of the category 2 compounds listed herein have been shown to turn on Nrf2, which turns on a grouping of anti-oxidant enzymes. Nrf2 also acts to reduce NF-K0 which reduces oxidation. Once Nrf2 is turned on, the compounds are not needed again until Nrf2 turns off, which usually takes a few days. Accordingly, the category 2 compounds should be taken in moderation. The dosing concentration and schedule disclosed herein reflects the moderation that should be used with the category 2 compounds.

[0046] The category 2 compounds in combination with category 1 compounds may act synergistically. The category 2 compounds in combination with category 1 compounds may balance methylation and oxidative signaling. The balancing of oxidative stress and methylation may optimize gene expression or overall patterns of gene expression. For instance, increased control and balance of oxidative stress may increase gene expression by reducing instances of DNA damage. Additionally, the composition may help regulate stability at the cellular level.

[0047] Compounds that regulate oxidative stress may produce a measurable effect on subjects. For example, multiple biomarkers may be observed to beneficially affect a subject. Biomarkers such as IL-6, IL-i , tumor necrosis factor-alpha, and C-reactive protein may be measured. The combination of category 1 and category 2 compounds may influence these biomarkers. In another example, the combined effect of category 1 and category 2 compounds may also beneficially affect gene expression patterns or cellular functions. The composition effects may be measurable through the above-listed biomarkers as well as through other mean. At the cellular level, the category 2 compounds may affect gene expression and oxidative response in a way that complements the effects of category 1 compounds.

[0048] Iron is an essential element in the cell and required for producing ATP in the mitochondria. While hemoglobin is the most important iron storage depot in humans, proteins like ferritin and transferrin are extremely important iron storage depots within the cell. Ferritin can be damaged with age resulting in a release of free iron which can react with important cell structures, and eventually lead to cell death. As such, the production of new ferritin to captureand bind to free iron is important to reducing aging-related damage and cell death related to free iron.

[0049] Compounds which promote the formation of ferritin, such as threonine, have been found to help prevent the ferritin-dependent “ferroptosis” which is a protective form of regulated cell death conceptually similar to apoptosis. As such, the ingestion of threonine helps to increase the lifespan of organisms by preventing the damage cause by ferroptosis and decreasing the amount of free iron within cells.

[0050] In one embodiment, the composition is administered in conjunction with fasting, wherein the subject undergoes a water-only fasting period before and during administration of the composition. In one method, the subject fasts for three days prior to administration, followed by two days of composition dosing. Fasting has been observed to result in changes in RNA expression after three days. In embodiments, the subject fasts during administration. In some embodiments, the subject fasts before administration and stops fasting during administration of the composition. In embodiments, the subject begins fast during administration.

[0051] In one embodiment, the composition is administered with proton pump inhibitors (PPIs) to reduce gastric discomfort associated with administration. The composition may cause acid reflux or stomach irritation, and co-admini strati on of PPIs before or during treatment may improve tolerability. This approach is particularly useful for subjects experiencing acid-related side effects during administration.

[0052] In one embodiment, the composition is formulated as a capsule for oral administration. The capsule may contain category 1 and category 2 compounds in specified ratios and may be enteric-coated to improve absorption and reduce gastric irritation. It may be designed for immediate-release, sustained-release, or delayed-release to optimize bioavailability.

[0053] In one embodiment, the composition is administered using an hourly dosing regimen to maintain a steady biological effect while reducing peak concentrations. Instead of four doses per day spaced approximately every five hours, the composition is administered every hour while awake, resulting in approximately sixteen doses per day. The total daily dose remainsunchanged, but each individual dose is proportionally reduced to 1 / 16th of the total daily amount.

[0054] For category 1 compounds, including methionine, S-adenosyl-methionine (SAM), betaine, glycine, serine, and threonine, the optimal dosage per administration may be about 0.00625 mmole / kg of subject, 0.0125 mmole / kg, 0.025 mmole / kg, 0.0375 mmole / kg, 0.05 mmole / kg, or 0.0625 mmole / kg.

[0055] For category 2 compounds, such as spermidine, taurine, cysteine, a-ketoglutarate, and hydrogen sulfide donors, the optimal dosage per administration is 0.00000625 moles / kg, 0.0000125 moles / kg, 0.000025 moles / kg, 0.0000375 moles / kg, 0.00005 moles / kg, or 0.0000625 moles / kg.

[0056] The hourly dosing approach provides a more consistent exposure profile, reducing fluctuations in compound concentration while maintaining sustained biological activity. This method may also mitigate gastrointestinal side effects, such as acid reflux, which has been noted as a potential concern.

[0057] Referring to Fig. 1, Fig. 1 is an illustration 100 of a pathway connection showing the link between category 1 compounds and category 2 compounds. The illustration 100 has three branches: a first branch 105, a second branch 110, and a third branch 115. The second branch 110 illustrates the methylation pathway beginning with methionine, which produces compounds that regulate oxidative stress in a subject. Methionine may form S-adenosylmethionine, which is converted to S-adenosylhomocysteine, then homocysteine. Homocysteine may subsequently form cysteine. Cysteine participates in oxidative stress regulation. It may produce hydrogen sulfide and may be utilized in protein synthesis. Cysteine may further react to form y- glutamylcysteine, which is subsequently converted to glutathione, a primary regulator of oxidative stress. Additionally, cysteine may convert into cysteine sulfonate and taurine, which also help regulate oxidative stress in cells.

[0058] The category 1 compounds listed herein support methylation pathways through the production of methionine. The first branch 105 and the third branch 115 illustrate methylation interactions that may result in the production of methionine. Many of the category 1 compoundslisted herein may participate in either the first branch 105 or the third branch 1 15, facilitating remethylation of homocysteine into methionine. Some category 1 compounds function specifically by donating methyl groups directly to homocysteine.

[0059] In the illustration shown in the second branch 110, the primary methyl donor is S- Adenosylmethionine. S-Adenosylmethionine (SAM), donates a methyl group to produce homocysteine, and the second branch branches from there to produce various compounds that treat oxidative stress in a subject.

[0060] The interaction between the second branch 110 and the third branch 115 illustrates the methylation cycle involving methylation and remethylation. Methionine may produce homocysteine by converting into S-adenosylmethionine and subsequently S- adenosylhomocysteine. Remethylation may occur through conversion of homocysteine back to methionine. For example, homocysteine may be remethylated back to methionine via methyl groups provided by betaine, which is derived from choline.

[0061] The interaction between the second branch 110 and the third branch 115 illustrates the methylation cycle involving methylation and remethylation. Methionine may produce homocysteine by converting into S-adenosylmethionine and subsequently S- adenosylhomocysteine. Remethylation may occur through the conversion of homocysteine back into methionine.

[0062] The terminal endpoints of the second branch 110 illustrate compounds resulting from these methylation pathways. These endpoints include glutathione and taurine, both of which regulate or reduce oxidative stress. Glutathione functions as an antioxidant that neutralizes oxidative cellular stress. Taurine regulates cellular stability and influences oxidative stress indirectly by supporting antioxidant defenses and scavenging reactive oxygen species.

[0063] The interaction between the second branch 110 and third branch 115 illustrates the methylation cycle involving methylation and remethylation, category 1 compounds generally support the methylation processes depicted, directly or indirectly producing category 2 compounds, category 2 compounds are shown at the endpoints or intermediate points of the second branch 110. category 2 compounds, such as glutathione and taurine, regulate oxidativestress and result from methylation pathways enabled by category 1 compounds. Accordingly, administering both category 1 and category 2 compounds to a subject may balance methylation and oxidative stress pathways at the cellular level, producing complementary effects.

[0064] Referring to Fig. 2, Fig. 2 is a biological age test result 200 subsequent to administration of an embodiment of the disclosed composition on a subject. This test quantifies the epigenetic age of multiple tissues based on the methylation status of related cytosines. Depending on which cytosines are methylated, and which cytosines are unmethylated, the epigenetic age of a tissue can be determined, and the overall epigenetic age of an individual can be determined as seen in age test result 200. For example, the subject in this test had an overall epigenetic age of 35.8, while his lung tissue had an epigenetic age of 25.9 based on the methylation status of various cytosines in his DNA. The subject also had a metabolic age of 38.1, a musculoskeletal age of 42.4, a blood age of 48.8, a liver age of 41.5, an inflammatory system age of 39.9, a kidney age of 45.3, a heart age of 35.8, a hormone system age of 39.8, an immune system age of 38.7 and a brain age of 40.9. These administration and result disclosed in the example below which led to these results is not intended to be inclusive of all aspects of the subject matter disclosed herein, but rather to illustrate representative methods, compositions, and results. These examples are not intended to exclude equivalents and variations of the present invention, which are apparent to one skilled in the art.

[0065] A 66 year old Caucasian male (subject) was treated with a composition of methionine, glycine, spermidine, cysteine and threonine. The subject was administered the composition four times a day, five hours apart for four consecutive days. The subject was given this treatment four times over a year.Composition of Dose

[0066] The subject was administered a dose of the following compounds: methionine, glycine, spermidine, threonine, and cysteine.

[0067] Methionine: Methionine is a “GRAS” compound, which is allowed for human consumption by the FDA. Methionine is the ingestible and absorbable precursor of S-Adenosyl methionine (SAM) which is the evolutionary designated singular (thus important decision point)methylating agent in the cell of DNA, RNA, and Proteins. One of the most significant items controlled by DNA methylation is the endogenous retrovirus LINE-1, which is necessary for development from the 2 to 8 cell stage (so multicellular life itself) but if active due to demethylation in late stages of life leads to the active virus in the cytoplasm which causes senescence. A re-zeroing of age happens at week three (showing evolution can re-zero cells) cleaning up the damage LINE-1 can cause at a time after which the germ cell line splits off from the somatic cell line in development. The somatic cell line’s amount of energy for repair damage will then be controlled (decreased) by a “DREAM” complex by active participation of information from the germ line cells. This throttling of damage repair has been seen as a key to the increase of somatic cell damage leading to aging.

[0068] Glycine: Glycine is the simplest amino acid and has been shown by the NIA’s ITP to be an agent (one of the few so far) that significantly increases the median life span of mice. It is a component for the synthesis of glutathione (the major small molecule antioxidant of the cell) which is the key to combating oxidation which has been seen by many as a key to damage leading to aging. Glycine was a GRAS compound until it was seen to have effects at higher doses in animal model organisms. It is now listed in 251 trials as a part of therapies for benefit by clinicaltrials.gov.

[0069] Spermidine: Spermidine is singular (thus important decision site) in biology in that it donates an amino-butyl group for the synthesis of hypusine at a specific lysine residue of the eukaryotic translation initiation factor 5A. Both Spermidine and hypusine have been seen to go down with age. Hypusine is important in making mitochondrial proteins (mitochondrial function goes down with age) and ATG3 needed for the induction of autophagy (needed to get rid of the damage of age). Spermidine also acts as a “sink” for the acyl groups taken off by Sirtuin enzymes that are needed to repair the increasing damage of age. Spermidine can also create hydrogen peroxide when needed which is the fundamental weak oxidant that other stronger oxidants are broken down into prior to be made into water. It is this hydrogen peroxide (quantity) that is assessed by the cell to determine the amount of oxidative attack it is under (since it becomes a sum of most all oxidants by this process) and if and when it is sufficient to turn on Nrf2 (transcription factor), this then creates all the antioxidant enzymes which in turn also turns off NF-KP (oxidation) and in turn also turns off CD-38 (only active with oxidation) which candegrade NAD+ and NMN which are needed by the Sirtuin and PARP enzymes which are involved with repair. Spermidine is listed as a therapy in 15 studies on clinicaltrials.gov.

[0070] Threonine: Threonine is a GRAS compound. It is a component that can be used in the synthesis of Methionine as well as up-regulating (thus increasing) the ferritin protein. With age and senescence the iron storage capacity of ferritin is compromised, this means more iron escapes from ferritin and then more iron is available to (exponentially) increase oxidation (thus decrease the quantity of glutathione which is the major small molecule antioxidant). Generally more oxidation leads to more damage which leads to more age. Threonine inhibits the ferritindependent “ferroptosis” which is a protective form of regulated cell death conceptually similar to apoptosis. Non-heme iron often increases in the brain with aging (as measured by MRI) and this increase correlates to decreases in cognitive performance.

[0071] Cysteine: Cysteine is a GRAS compound. It is a component in the creation of Glutathione mentioned above with Glycine also mentioned above. Its availability also helps the methionine cycle rotate to make methionine instead of having to make cysteine from homocysteine.

[0072] The amounts of each composition were set so that by the time the subject was administered the composition for the fourth time, a final dosage of approximately 0.5 x 10-3 methionine moles / kg body weight per dose, 0.5 x 10-3 glycine moles / kg body weight per dose, 1 x 10-4 spermidine moles / kg body weight per dose, 1 x 10-4 cysteine moles / kg body weight per dose and 0.5 x 10-3 threonine moles / kg body weight per dose was given to the subject.

[0073] In addition to the test result 200, the subject has his Epigenetic age periodically tested using DNA methylation-based biological clock tests provided by myDNAge, Clock GRIM Age, TruMeLabs and Blueprint Biomarkers. The testing periods and the resulting Epigenetic age results are presented in Table 1 below.Table 1

[0074] Based on these results, there appears to be an observed decrease in DNA methylation after the administration of the procedure, with long term decreases that occur after the compounds have been taken. This can be seen in the decrease from 55.50 to 54.40 between August 12, 2024 and October 31, 2024.

[0075] In another embodiment, dosages are provided three times per day, once every six hours.The example dosages of methione, glycine, threonine, spermidine and cystine are provided in Table 2 below.Tab e 2

[0076] Referring to Fig. 3, Fig. 3 an illustration 300 of an administration timeline in an embodiment of the disclosed subject matter. In various embodiments, the subject is dosed periodically over a defined period of time. For instance, the subject may be dosed in pulsesseparated by intervals ranging from days to weeks or even months. In one example, a subject may be administered four pulses within a year.

[0077] Each pulse may include a dosing period during which the subject receives doses at regular intervals. For example, during a pulse, the subject may receive a dose every hour. In embodiments, the dosing intervals may vary from once per day to once per hour. In one example, a subject may be administered a dose once every four hours. In each of these cases, doses typically would not be administered while the subject sleeps, for instance, during an 8-hour sleep period per day.

[0078] As shown in illustration 300, each pulse may include additional time periods in addition to the periodic dosing period 310. For example, a pulse may include a fasting period 305, where the subject fasts for a period of time prior to the dosing period 310. In embodiments, the fasting period may include a water fast, where the subject limits food intake to water or low-calorie foods. In embodiments, the fast may include consuming a restricted-calorie diet. For instance, the subject may be limited to half the normal daily calorie intake during the fasting period 305. In embodiments, the fasting period may limit the subject to approximately one-quarter of the normal daily calorie intake appropriate for their body size.

[0079] In embodiments, such as the embodiment shown in illustration 300, a pulse may include a cessation period 315, during which the subject is not administered any doses and does not fast. The cessation period may last various lengths of time, such as between one day and one year. In embodiments, the cessation period may continue until the beginning of the next pulse. In embodiments, the subject may undergo multiple pulses per year. For instance, between one and ten pulses may be administered to a subject over the course of a year. In various tests, the beneficial results are observed during the cessation period when the subject refeeds.

[0080] In embodiments, the subject continues to fast during the dosing period. This is so the therapy has no effects from diet.

[0081] Category 1 Compounds

[0082] Category 1 compounds include: S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin Bl 2, glycine, serine, threonine.

[0083] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes S-5’-adenosyl-L -methionine (SAM). SAM serves as a key methyl donor in cellular methylation. It transfers methyl groups to DNA, RNA, proteins, and small molecules to regulate gene expression, protein function, and metabolic pathways.

[0084] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes methionine. Methionine is an essential amino acid that provides methyl groups for DNA and protein methylation. It supports cellular function and gene expression.

[0085] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes betaine. Betaine donates methyl groups to convert homocysteine to methionine. This process aids DNA methylation and maintains cellular methylation balance.

[0086] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes choline. Choline oxidizes to betaine and supplies methyl groups for DNA methylation. It also supports gene expression and cellular function.

[0087] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes folate. Folate in the form of tetrahydrofolate (THF) transfers single-carbon units. This enables nucleic acid metabolism, amino acid synthesis, DNA methylation, and normal cell division.

[0088] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes vitamin B12. Vitamin B12 assists methionine synthase in remethylating homocysteine to methionine. It contributes to DNA methylation and overall cellular metabolism.

[0089] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes glycine. Glycine contributes to one-carbon metabolism by supplying methyl groups for DNA methylation and biomolecule synthesis.

[0090] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes serine. Serine supports folate-driven one-carbon metabolism. It provides carbon units for methyl donors to aid DNA methylation and gene regulation.

[0091] In a specific example, the category 1 compound used in the composition of the disclosed subject matter includes threonine. Threonine metabolizes into glycine and contributes to the one-carbon cycle. This supplies methyl groups for DNA and protein methylation.

[0092] Category 2 Compounds

[0093] Category 2 compounds include: spermidine, spermine, calcium peroxide, zinc, N- acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, O2* , OH*, ’O2, O3, ROOH, Na2S, H>O2, ROS, RNS, RCS, RSOH, O2’, O2, H2S, O3, HOC1, HOBr, HOI, ROOM (where R is alkyl, cycloalkyl, heteroalkyl, heterocycloalkyl, alkenyl, heteroalkenyl, cycloalkenyl, or heterocycloalkenyl), H2O2generators (e.g., metformin, acetaminophen), ortho-hydroxyphenols oxidizable to quinones, para-dihydroxyphenols oxidizable to quinones, quinones (oxidized aromatic derivatives), hydrogen sulfide (H2S), H2S donors (e.g., NaHS, Na2S, diallyl trisulfide (DATS), GYY4137 (patent # WO2014018569 Al), SG-1002 (SulfaGENEX), penicillamine-based donors, polyorganosulfides, 2-mercaptoethanol, dithiothreitol, isothiocyanates, sulforaphane (broccoli), glucoraphanin (broccoli), curcumin (turmeric), pyrrolidone (water-soluble), Theracurmin (nanoparticle), zerumbone, cinnamate analogs (e.g., cinnamic aldehyde), quercetin (onions, apples, tea), isoquercetin (enhanced absorption), kaempferol, ginseng (Panax ginseng, Panax quinquefolius), carnosic acid, xanthohumol, Dh404, (R)-alpha-lipoic acid, benzyl isothiocyanate, neoglucobrassicin, glucosinolates, hydrophilic oxidized lycopene derivatives, 4-hydroxynonenal (HNE), 15- deoxydelta prostaglandin J2 (15-dPGJ2), falcarindiol, hydroxytyrosol, barley beta-glucan, luteolin, pyrroloquinoline quinone, mangafodipir trisodium (MnDPDP), ATB-346 (Antibe Therapeutics), NBS-1120 (City College of New York), GIC-101 (GI Care Pharma), AP39 (patent # WO2013045951A1, University of Exeter), AP67, AP97, AP105 (WO2014018569A1), Sialor, Sulfarlem, Anetholetrithione, DHEA, coal tar, garlic (via H2S), -lapachone (South American tree bark), pterostilbene, apigenin (parsley), l,4-diphenyl-l,2,3-triazoles, 15-deoxy- A12,14-prostaglandin J2, 3,4-dihydroxyphenylethanol, 3-alkylamino-lH-indole acrylates, 4-phenyl-l,2,4-triazole derivatives, 6-shogaol (ginger), acetyl-1 l-keto-0-boswellic acid, acteoside, allicin, astaxanthin, bardoxolone (CDDO), methyl ester of bardoxolone (CDDO-Me), benfotiamine, butein, tert-butylhydroquinone (tBHQ), caffeine, cardamonin, camosol, catechin, cinnamic acid derivatives (e.g., caffeic acid phenethyl ester, ferulic acid ethyl ester, trans- cinnamaldehyde), curcumin derivatives / analogues, dimeric ferulic acid derivative, dimethyl fumarate (DMF, Tecfidera), diterpenoid derivatives, epicatechin, epigallocatechin, green tea, epigallocatechingallate (EGCG), eriodictyol-7-O-glucoside, fisetin, genistein, licochalcone E, naphthazarin, naringenin, nordihydroguaiaretic acid (NDGA), phenethyl isothiocyanate (PEITC), phloretin, piperlongumine and analogs, pyrrolidine dithiocarbamate, quercetin, resveratrol, silybin, alpha (a), beta (0), gamma (y), delta (5) tocotrienols (T3), triptolide, fenofibrate, melatonin, vinblastine, artichokes, cyanoside-3 -O-glucoside (anthocyanins), cocoa, FW1256 and compounds 1-51 (Li et al ., European Journal of Medicinal Chemistry, 07 Nov 2019, 185: 111862), methylene blue, and optionally a carrier, or any combination thereof.

[0094] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes spermidine. Spermidine is a polyamine present in all living cells and counteracts oxidative damage. It enhances autophagy to reduce cellular stress.

[0095] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes spermine. Spermine is a polyamine that protect cells from oxidative damage. It stabilizes DNA and regulates ion channels.

[0096] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes calcium peroxide. Calcium peroxide releases hydrogen peroxide upon decomposition in water which then turns on Nrf2 to modulate oxidative stress.

[0097] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes zinc. Zinc serves as a cofactor for antioxidant enzymes. It stabilizes cell membranes and prevents lipid peroxidation to maintain cellular health.

[0098] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes N-acetylcysteine. N-acetylcysteine boosts glutathioneproduction to reduce oxidative stress. It acts as a mucolytic and counters acetaminophen overdose.

[0099] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cysteine. Cysteine supports glutathione synthesis to counteract oxidative stress. Its sulfur content aids detoxification and protein structure through disulfide bonds.

[0100] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes a-ketoglutarate. a-ketoglutarate in the TCA cycle reduces reactive oxygen species. It serves as a substrate for amino acid metabolism.

[0101] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes fucoidans. Fucoidans from brown seaweed reduce oxidative stress with antioxidant properties. Fucoidans have been shown to turn on Sirtuin 6. They also exhibit anti-inflammatory and anticancer effects.

[0102] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes harmine. Harmine is a [3-carboline alkaloid with antioxidant properties. It’s studied for neuroprotection and oxidative stress regulation.

[0103] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes taurine. Taurine turns on Nrf2 to protect cells. It regulates calcium levels and osmoregulation.

[0104] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes ergothioneine. Ergothioneine neutralizes reactive oxygen species to reduce oxidative stress. It accumulates in high-stress tissues like liver and kidneys.

[0105] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes urolithin A. Urolithin A from ellagitannin triggers mitophagy to decrease oxidative stress. It improves mitochondrial function for cellular health.

[0106] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes terpenoids. Terpenoids from plants reduce oxidative stress through antioxidant effects. They also provide anti-inflammatory and anticancer benefits.

[0107] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes berberine. Berberine is an isoquinoline alkaloid that reduces oxidative stress with antioxidant activity. It supports metabolic and cardiovascular health.

[0108] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes superoxide anion (O?* ). Superoxide anion is a reactive oxygen species that can damage cells. Antioxidant enzymes like SOD convert it to less reactive forms.

[0109] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes ozone (Os). Ozone acts as a strong oxidizer at ground level and increases oxidative stress. Antioxidants mitigate its effects on respiratory tissues.

[0110] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydroperoxides (ROOH). Hydroperoxides with a peroxide group decompose into free radicals. This elevates oxidative stress and potential cellular damage.[0U1] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes sodium sulfide (Na2S). Sodium sulfide releases ILS with antioxidant properties. This may decrease oxidative stress.

[0112] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrogen peroxide (H2O2). Hydrogen peroxide is a reactive oxygen species that can damage cells. It also functions as a signaling molecule in biological processes.

[0113] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes reactive oxygen species (ROS). Reactive oxygen species are oxygen-containing molecules that damage DNA, proteins, and lipids. This disrupts cellular function.

[0114] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes reactive nitrogen species (RNS). Reactive nitrogen species like peroxynitrite induce nitrosative stress. This damages cells and contributes to disease.

[0115] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes reactive carbonyl species (RCS). Reactive carbonyl species modify proteins and nucleic acids. This leads to cellular dysfunction and oxidative stress.

[0116] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes sulfenic acids (RSOH). Sulfenic acids from thiol oxidation participate in redox signaling. They trigger oxidative stress responses.

[0117] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes molecular oxygen (O2). Molecular oxygen essential for respiration can form reactive oxygen species. This contributes to oxidative stress.

[0118] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrogen sulfide (H2S). Hydrogen sulfide is a signaling gas with antioxidant effects. It reduces oxidative stress in biological systems.

[0119] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hypochi orous acid (HOC1). Hypochi orous acid produced by immune cells oxidizes strongly. It destroys pathogens but may harm host tissues.

[0120] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hypobromous acid (HOBr). Hypobromous acid is a reactive species that oxidizes biomolecules. This increases oxidative stress and cellular damage.

[0121] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hypoiodous acid (HOI). Hypoiodous acid is an iodine-based species that modifies proteins and lipids. It contributes to oxidative stress.

[0122] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes alkyl hydroperoxides (ROOM). Alkyl hydroperoxides withorganic groups form free radicals upon decomposition. This raises oxidative stress and cellular damage.

[0123] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrogen peroxide (H2O2) generators, such as metformin or acetaminophen. Hydrogen peroxide generators like metformin or acetaminophen produce H2O2 in cells. This may increase oxidative stress or trigger antioxidant responses.

[0124] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes ortho-hydroxy phenols that can be oxidized to quinones. Orthohydroxyphenols oxidize into quinones. These undergo redox cycling and generate reactive oxygen species to elevate oxidative stress.

[0125] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes para-dihydroxyphenols that can be oxidized to quinones. Paradihydroxyphenols form quinones upon oxidation. This promotes redox cycling and increases oxidative stress.

[0126] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes quinones. Quinones from oxidized aromatics undergo redox cycling. They produce reactive oxygen species and increase oxidative stress.

[0127] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrogen sulfide (H2S). Hydrogen sulfide serves as a gaseous signaling molecule with antioxidant properties. It reduces oxidative stress in biological systems.

[0128] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrogen sulfide (H2S) donors, such as sodium hydrosulfide (NaHS), sodium sulfide (NaiS), diallyl trisulfide (DATS), GYY4137, SG-1002, and penicillamine-based H2S donors. Hydrogen sulfide donors like NaHS and DATS release ILS with antioxidant effects. This reduces oxidative stress.

[0129] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes polyorganosulfides. Polyorganosulfides release ILS with antioxidant properties. This decreases oxidative stress in biological systems.

[0130] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 2-mercaptoethanol. 2-mercaptoethanol is a thiol compound that reduces disulfide bonds in proteins. This alters their structure and function.

[0131] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes dithiothreitol. Dithiothreitol is a strong reducing agent that cleaves protein disulfide bonds. It impacts cellular processes.

[0132] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes isothiocyanates. Isothiocyanates naturally occurring induce phase II detoxification enzymes. This enhances defenses against oxidative damage.

[0133] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes sulforaphane. Sulforaphane from broccoli activates antioxidant response elements. It strengthens cellular resilience against oxidative stress.

[0134] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes glucoraphanin. Glucoraphanin from broccoli is a precursor to sulforaphane and supports antioxidant defenses. It neutralizes reactive oxygen species.

[0135] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes curcumin. Curcumin from turmeric maintains cellular integrity under oxidative conditions.

[0136] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes water-soluble pyrrolidone. Water-soluble pyrrolidone acts as a precursor to antioxidants. It helps reduce oxidative stress.

[0137] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes theracurmin. Theracurmin is a nanoparticle form of curcumin with enhanced bioavailability. It reduces oxidative stress effectively.

[0138] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes zerumbone. Zerumbone from tropical ginger modulates oxidative stress pathways. It maintains cellular redox balance.

[0139] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cinnamic aldehyde. Cinnamic aldehyde increases glutathione levels and activates Nrf2. This reduces oxidative stress.

[0140] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes quercetin. Quercetin scavenges reactive oxygen species and prevents lipid peroxidation. It decreases oxidative stress.

[0141] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes isoquercetin. Isoquercetin boosts glutathione production and reduces oxidative DNA damage. This lowers stress levels.

[0142] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes kaempferol. Kaempferol reduces reactive oxygen species and suppresses inflammation. It decreases oxidative stress.

[0143] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes ginseng. Ginseng stabilizes mitochondrial function and reduces reactive oxygen species. It enhances antioxidant enzyme activity.

[0144] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes carnosic acid. Carnosic acid scavenges free radicals and activates Nrf2. It prevents lipid peroxidation and reduces oxidative stress.

[0145] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes xanthohumol. Xanthohumol inhibits NADPH oxidase and increases antioxidant protein expression. This lowers oxidative stress.

[0146] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes Dh404. Dh404 activates Nrf2 and boosts detox enzyme expression. It prevents oxidative damage.

[0147] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes (R)-alpha-lipoic acid. (R)-alpha-lipoic acid scavenges reactive oxygen species and regenerates glutathione, vitamin C, and E. It reduces oxidative stress.

[0148] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes isothiocyanate. Isothiocyanate activates Nr£2 and increases antioxidant enzyme production. This decreases oxidative stress.

[0149] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes benzyl isothiocyanate. Benzyl isothiocyanate triggers Nrf2 and raises glutathione levels. It reduces oxidative stress.

[0150] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes neoglucobrassicin. Neoglucobrassicin induces detoxification enzymes and modulates Nrf2. This lowers oxidative stress.

[0151] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes glucosinolates. Glucosinolates form isothiocyanates and activate Nrf2. They increase antioxidant enzyme levels to reduce stress.

[0152] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydrophilic oxidized derivatives of lycopene. Hydrophilic oxidized derivatives of lycopene scavenge free radicals and stabilize mitochondria. They decrease oxidative stress.

[0153] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 4-hydroxynonenal (HNE). 4-hydroxynonenal at low levels induces antioxidant enzymes to reduce stress. At high levels, it damages proteins, DNA, and lipids.

[0154] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 15-deoxydelta prostaglandin J2 (15-dPGJ2). 15-deoxydelta prostaglandin J2 activates PPAR-y and Nrf2 to boost detox enzyme expression. It reduces oxidative stress.

[0155] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes falcarindiol. Falcarindiol curbs reactive oxygen species production and suppresses inflammation. It decreases oxidative stress.

[0156] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes hydroxytyrosol. Hydroxytyrosol increases glutathione levels. It reduces oxidative stress.

[0157] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes barley beta-glucan. Barley beta-glucan enhances antioxidant enzyme activity. It reduces inflammation and stress.

[0158] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes luteolin. Luteolin activates Nrf2. It prevents lipid peroxidation and reduces stress.

[0159] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes pyrroloquinoline quinone. Pyrroloquinoline quinone boosts mitochondrial biogenesis. It activates Nrf2 to decrease stress.

[0160] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes mangafodipir trisodium (MnDPDP). Mangafodipir trisodium offers antioxidant properties. It reduces oxidative stress.

[0161] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes garlic. Garlic produces ILS. It prevents lipid peroxidation and reduces stress.

[0162] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes P-lapachone. P-lapachone cycles NADH to NAD+to balance redox.

[0163] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes pterostilbene. Pterostilbene activates Nrf2. It boosts antioxidant enzymes to reduce stress.

[0164] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes apigenin. Apigenin activates Nrf2. It prevents lipid peroxidation and reduces stress.

[0165] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes l,4-diphenyl-l,2,3-triazole. 1,4-diphenyl- 1,2, 3 -triazole stabilizes redox balance. It decreases oxidative stress.

[0166] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 15-deoxy-A12,14-prostaglandin J2. 15-deoxy-A12,14- prostaglandin J2 is a PGD2 metabolite that binds PPARy. It promotes adipocyte differentiation and reduces inflammation.

[0167] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 3,4-dihydroxyphenylethanol. 3,4-dihydroxyphenylethanol is a phenolic from olive oil that enhances antioxidant effects. It offers anti-inflammatory and neuroprotective benefits.

[0168] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 3-alkylamino-lH-indole acrylates. 3-alkylamino-lH-indole acrylates activate Nrf2. They support cellular defense against oxidative stress.

[0169] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 4-phenyl-l,2,4-triazole derivatives. 4-phenyl-l,2,4-triazole derivatives like PTAD are reactive dienophiles used in organic synthesis. They derive from 4- phenylurazole oxidation.

[0170] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes 6-shogaol (from ginger). 6-shogaol from ginger reduces inflammation and oxidative stress. It supports gut health and chronic disease prevention.

[0171] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes acetyl- 11-keto-P-boswellic acid. Acetyl-11-keto-P-boswellic acid from frankincense curbs inflammation. It may aid arthritis treatment.

[0172] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes acteoside. Acteoside is a plant glycoside with antioxidant and anti-inflammatory effects. It supports brain health and reduces oxidative stress.

[0173] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes allicin. Allicin from garlic offers antibacterial and antiviral properties. It also supports cardiovascular health.

[0174] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes astaxanthin. Astaxanthin is a carotenoid that protects cells from oxidative stress. It improves skin health and reduces inflammation.

[0175] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes bardoxolone (CDDO). Bardox olone is a synthetic triterpenoid that activates Nrf2. It reduces inflammation and oxidative stress in kidney disease.

[0176] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes methyl ester of bardoxolone (CDDO-Me). Methyl ester of bardoxolone is a bioavailable form that activates Nrf2. It targets oxidative stress in chronic kidney disease.

[0177] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes benfotiamine. Benfotiamine is a fat-soluble Bl form that enhances nerve function. It reduces oxidative stress in diabetic neuropathy.

[0178] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes butein. Butein is a plant flavonoid that reduces oxidative stress and inflammation. It supports cardiovascular health.

[0179] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes tert-butylhydroquinone (tBHQ). Tert-butylhydroquinone is a synthetic antioxidant used in food preservation. It protects cells from oxidative damage, though safety remains debated.

[0180] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes caffeine. Caffeine is a stimulant that increases alertness. It enhances physical and cognitive function.

[0181] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cardamonin. Cardamonin from cardamom reduces inflammation and oxidative stress. It may prevent chronic diseases.

[0182] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes carnosol. Carnosol from rosemary provides antioxidant effects. It supports brain health and reduces oxidative stress.

[0183] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes catechin. Catechin from tea and fruits improves heart health and metabolism.

[0184] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cinnamic acid and its derivatives (e.g., caffeic acid phenethyl ester, ferulic acid ethyl ester, and trans-cinnamaldehyde). Cinnamic acid and its derivatives reduce oxidative stress and inflammation. They support cardiovascular health and wellness.

[0185] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes curcumin and its derivatives and analogues. Curcumin and its derivatives reduce inflammation and oxidative stress. They support brain health and chronic disease prevention.

[0186] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes dimeric derivative of ferulic acid. Dimeric derivative of ferulic acid enhances antioxidant effects. It supports heart health and reduces oxidative stress.

[0187] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes dimethyl fumarate (DMF, Tecfidera). Dimethyl fumarate activates Nrf2 to reduce inflammation and oxidative stress. It treats multiple sclerosis.

[0188] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes diterpenoid derivatives. Diterpenoid derivatives reduce inflammation and exhibit anticancer effects. They offer therapeutic potential for various diseases.

[0189] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes epicatechin. Epicatechin from chocolate and tea improves heart and brain health.

[0190] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes epigallocatechin. Epigallocatechin from green tea reduces oxidative stress. It supports heart health and metabolism.

[0191] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes green tea. Green tea rich in catechins provides antioxidant effects. It enhances heart and brain health.

[0192] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes epigallocatechingallate (EGCG). Epigallocatechingallate from green tea reduces inflammation and oxidative stress. It supports heart health and weight loss.

[0193] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes eriodictyol-7-O-glucoside. Eriodictyol-7-O-glucoside is a flavonoid that reduces oxidative stress. It supports heart health and curbs inflammation.

[0194] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes fisetin. Fisetin from strawberries enhances antioxidant effects. It improves cognition and reduces age-related disease.

[0195] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes genistein. Genistein from soybeans provides antioxidant and estrogenic effects. It supports heart health and hormone balance.

[0196] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes licochalcone E. Licochalcone E from licorice reduces oxidative stress. It enhances skin and metabolic health.

[0197] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes naphthazarin. Naphthazarin is an anthraquinone with antioxidant effects. It may inhibit cancer cell growth.

[0198] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes naringenin. Naringenin from citrus reduces oxidative stress. It supports metabolism and heart health.

[0199] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes nordihydroguaiaretic acid (NDGA). Nordihydroguaiaretic acid is a lignan that reduces inflammation and oxidative stress. It may aid cancer and heart disease treatment.

[0200] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes phenethyl isothiocyanate (PEITC). Phenethyl isothiocyanate from cruciferous vegetables reduces cancer risk and oxidative stress. It inhibits tumor growth.

[0201] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes phloretin. Phloretin from apples reduces oxidative stress. It enhances skin health and insulin sensitivity.

[0202] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes piperlongumine and its analogs. Piperlongumine from piper longum reduces cancer risk and oxidative stress. It enhances cancer therapy efficacy.

[0203] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes pyrrolidine dithiocarbamate. Pyrrolidine dithiocarbamate inhibits NF-KB to reduce inflammation. It mitigates oxidative stress and aids cancer treatment.

[0204] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes quercetin. Quercetin from fruits and vegetables scavenges free radicals. It improves heart and immune health.

[0205] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes resveratrol. Resveratrol from grapes reduces oxidative stress. It supports heart health and longevity.

[0206] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes silybin. Silybin from milk thistle protects the liver. It reduces inflammation and oxidative stress.

[0207] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes alpha (a), beta ((B), gamma (y), and delta (8) tocotrienols (T3). Alpha, beta, gamma, and delta tocotrienols are vitamin E forms that reduce oxidative stress. They improve heart and brain health.

[0208] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes triptolide. Triptolide from Tripterygium reduces inflammation and cancer risk. It treats autoimmune and inflammatory conditions.

[0209] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes fenofibrate. Fenofibrate lowers lipids and improves heart health. It reduces cholesterol and triglycerides.

[0210] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes melatonin. Melatonin regulates sleep and reduces oxidative stress. It enhances immune function.

[0211] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes vinblastine. Vinblastine from periwinkle treats cancer. It halts cell division in tumors.

[0212] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes artichokes. Artichokes rich in antioxidants support liver health. They reduce cholesterol and oxidative stress.

[0213] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cyanoside-3 -O-glucoside (anthocyanins). Cyanoside-3-O- glucoside is an anthocyanin that reduces oxidative stress. It improves heart health and wellness.

[0214] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes cocoa. Cocoa high in flavonoids reduces oxidative stress. It supports heart health and mood.

[0215] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes methylene blue. Methylene blue activates nuclear factor erythroid 2-related factor 2 (NRF2) which is a transcription factor that regulates oxidative stress and toxic substances at a cellular level.

[0216] In a specific example, the category 2 compound used in the composition of the disclosed subject matter includes ginger. Ginger contains bioactive compounds that reduce oxidative stress. It controls inflammation and supports overall health.

[0217] A composition for administering to a subject includes a first compound chosen from S- 5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B 12, glycine, serine, threonine, and any combination thereof, and a second compound chosen from spermidine, spermine, calcium peroxide, zinc, N-acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, ROOH, ROS, RNS, RCS, RSOH, ROOM, where R is alkyl, cycloalkyl, heteroalkyl, heterocycloalkyl, alkenyl, heteroalkenyl, cycloalkenyl, or heterocycloalkenyl, 021, 02, H2S, 03, H0C1, HOBr, HOI, Na2S, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaHS), sodium sulfide (Na2S), diallyl trisulfide (DATS), metformin, acetaminophen, ortho hydroxyphenols, para dihydroxyphenols, and optionally, a carrier, and any combination thereof. The composition may comprise methionine and spermidine. The composition may be formulated as an injectable solution, a tablet, or a capsule. The composition may be in an amount sufficient to beneficially change a surrogate marker for aging level in a human when compared to the surrogate marker level prior to administration, where the change may be a lowered aging level. The surrogate marker may be C-reactive protein, tumor necrosis factor-alpha, Interleukin-6 (IL-6), IL-1 , or DNA methylation, and the change may be correlated to health span and life span.

[0218] A method for reducing inflammation in a subject includes administering the composition to the subject. The composition may be administered at a dosage of at least 0.1 x 10A-3 to 1.0 x 10A-3 moles / kg of the first compound and 1 x 10A-6 moles / kg to 1 x 10A-3 moles / kg of the second compound. The administration may occur over four pulses per day, five hours apart, for four to six consecutive days. The composition may be ingested over four to six days, four times daily, and may be in an aerosol, lyophilized, powder, or emulsion form. The subject may be a human who receives the composition for at least four days but not more than four, five, or six days. The composition may be in a tablet or capsule administered orally at least four times daily. The composition may be administered to the subject four times daily for not more than four days.

[0219] A method for administering a compound to a subject for beneficially changing a surrogate marker for aging level includes administering a compound comprising a first compound chosen from S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B 12, serine, and any combination thereof, a second compound chosen from spermidine, spermine, calcium peroxide, zinc, N-acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, ROOH, ROS, RNS, RCS, RSOH, ROOM, where R is alkyl, cycloalkyl, heteroalkyl, heterocycloalkyl, alkenyl, heteroalkenyl, cycloalkenyl, or heterocycloalkenyl, 021, 02, H2S, 03, H0C1, HOBr, HOI, Na2S, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaHS), sodium sulfide (Na2S), diallyl trisulfide (DATS), metformin, acetaminophen, ortho hydroxyphenols, para dihydroxyphenols, and any combination thereof, and a third compound chosen from threonine, N-acetylcysteine, glycine, and any combination thereof. The compound may be administered at a dosage of at least 0.1 x 10A-3 to 1.0 x 10A-3 moles / kg of the first compound, 1 x 10A-6 moles / kg to 1 x 10A-3 moles / kg of the second compound, and at least 0.1 x 10A-3 to 1.0 x 10A-3 moles / kg of the third compound. The administration may occur over four pulses per day, five hours apart, for four to six consecutive days. The compound may be ingested over four to six days, four times a day, and may be in an aerosol, lyophilized, powder, or emulsion form. The subject may be a human who receives the compound for at least four days but not more than four, five, or six days. The compound may be administered in a tablet or capsule orally at least four times daily. The compound may be administered in a pulse preceded by a water fast, where the water fast may last about three days, the pulse may be between about one and two days, and the pulse mayinclude administering a dose multiple times per day, once every hour during the pulse. The dose may be administered in enteric capsules.

[0220] Many variations may be made to the embodiments described herein. All variations, including combinations of embodiments, are intended to be included within the scope of this disclosure. The description of the embodiments herein can be practiced in many ways. Any terminology used herein should not be construed as restricting the features or aspects of the disclosed subject matter. The scope should instead be construed in accordance with the appended claims.

Claims

CLAIMS:

1. A composition for administering to a subject, the composition comprising: a first compound chosen from: S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B12, glycine, serine, threonine, and any combination thereof; and a second compound chosen from: spermidine, spermine, calcium peroxide, zinc, N- Acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, 03, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaSH), sodium sulfide (Na2S), metformin, acetaminophen, curcumin, quercetin, ginseng, R-alpha-lipoic acid, apigenin, fisetin, melatonin, ginger, astaxanthin methylene blue and optionally, a carrier, and any combination thereof.

2. The composition of claim 1, wherein the first compound composition is methionine, betaine, or both.

3. The composition of claim 1, wherein the second compound composition is spermidine, Spermine, cysteine, taurine, H202, H2S, NaSH, or any combination thereof.

4. The composition of claim 1, wherein the first compound composition and second compound composition are in an amount sufficient to beneficially change a surrogate marker for aging level in a human when compared to the surrogate marker level prior to administration.

5. The composition of claim 4 wherein the change in the level of the surrogate marker is a lowered aging level.

6. The composition of claim 5, wherein the surrogate marker is C-reactive protein, tumor necrosis factor-alpha, Interleukin-6 (IL-6), or IL-1 .

7. The composition of claim 4, wherein the change in the level of the surrogate marker is correlated to health-span and or life-span.

8. The composition of claim 7, wherein the surrogate marker is DNA methylation.

9. The composition of claim 1, wherein the composition comprises methionine and spermidine.

10. An injectable formulation, comprising the composition of claim 1 .

11. A tablet or capsule comprising the composition of claim 1.

12. A method of reducing inflammation in a subject, comprising: administering to the subject the composition of claim 1.

13. The method of claim 12, wherein the composition is administered to a subject at a dosage of at least 0.1 x 10A-3 to 1.0 X 10A-3 moles / kg of the first compound to the subject, and1 x 10A-6 moles / kg to 1 x 10A-3 moles / kg of the second compound to the subject; wherein said administered compositions are given over four pulses per day, 5 hours apart, for 4 to 6 consecutive days total at component concentrations between 0.1 x 10A-3 and 1.0 x 10A-3 moles / kg of a weight of the subject is optimally given.

14. The method of claim 12, wherein the composition is ingested over 4-6 days, 4 times a day.

15. The method of claim 12, wherein the composition is in an aerosol, lyophilized, powder, or emulsion form.

16. The method of claim 12, wherein the subject is a human.

17. The method of claim 16, wherein the composition is administered to the human for at least 4 days.

18. The method of claim 16, wherein the composition is administered to the human for not more than 4, 5, or 6 days.

19. The method of claim 12, wherein the composition is in a tablet or capsule that is administered orally at least four times daily.

20. The method of claim 12, wherein the composition is administered to the subject four times daily for not more than 4 days.

21. The method of claim 12, wherein the composition comprises methionine and spermidine.

22. A composition for administering to a subject, the composition comprising:a first compound chosen from: S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B 12, serine, and any combination thereof; a second compound chosen from: spermidine, spermine, calcium peroxide, zinc, N- Acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, , H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaHS), sodium sulfide (Na2S), metformin, acetaminophen, curcumin, quercetin, ginseng, R-alpha-lipoic acid, apigenin, fisetin, methylene blue, melatonin, ginger, astaxanthin and optionally, a carrier, and any combination thereof; and a third compound chosen from: threonine, N-acetylcysteine, glycine, and any combination thereof.

23. The composition of claim 22, wherein the first compound composition, second compound composition and third compound composition are in an amount sufficient to beneficially change a surrogate marker for aging level in a human when compared to the surrogate marker level prior to administration.

24. The composition of claim 23 wherein the change in the level of the surrogate marker is a lowered aging level.

25. The composition of claim 24, wherein the surrogate marker is C-reactive protein, tumor necrosis factor-alpha, Interleukin-6 (IL-6), or IL-1 .

26. The composition of claim 23, wherein the change in the level of the surrogate marker is correlated to health-span and or life-span.

27. The composition of claim 26, wherein the surrogate marker is DNA methylation.

28. The composition of claim 22, wherein the composition comprises methionine and spermidine.

29. An injectable formulation, comprising the composition of claim 22.

30. A tablet or capsule comprising the composition of claim 22.

31. A method of administering a compound to a subject for beneficially changing a surrogate marker for aging level in a human when compared to the surrogate marker level prior to administration, wherein said method comprises:administering to a subject a compound; wherein said compound is comprised of: a first compound chosen from: S-5'-adenosyl-L-methionine (SAM), methionine, betaine, choline, folate, vitamin B 12, serine, and any combination thereof; a second compound chosen from: spermidine, spermine, calcium peroxide, zinc, N- Acetylcysteine, cysteine, a-ketoglutarate, fucoidans, harmine, taurine, ergothioneine, urolithin A, terpenoids, berberine, 03, Na2S, H2O2, hydrogen sulfide (H2S), sodium hydrosulfide (NaHS), sodium sulfide (Na2S), metformin, acetaminophen, curcumin, quercetin, ginseng, R-alpha-lipoic acid, apigenin, fisetin, methylene blue, melatonin, ginger, astaxanthin and optionally, a carrier, and any combination thereof; and a third compound chosen from: threonine, N-acetylcysteine, glycine, and any combination thereof; wherein the subject is administered a dosage of at least 0.1 x 10A-3 to 1.0 X 10A-3 moles / kg of the first compound to the subject, 1 x 10A-6 moles / kg to 1 x 10A-3 moles / kg of the second compound to the subject, and of at least 0.1 x 10A-3 to 1.0 X 10A-3 moles / kg of the third compound to the subject; wherein said administrations are given over four pulses per day, 5 hours apart, for 4 to 6 consecutive days total at component concentrations between 0.1 x 10A-3 and 1.0 x 10A-3 moles / kg of a weight of the subject is optimally given.

32. The method of claim 31, wherein the compound is ingested over 4-6 days, 4 times a day.

33. The method of claim 31, wherein the compound is in an aerosol, lyophilized, powder, or emulsion form.

34. The method of claim 31, wherein the subject is a human.

35. The method of claim 34, wherein the compound is administered to the human for at least4 days.

36. The method of claim 34, wherein the compound is administered to the human for not more than 4, 5, or 6 days.

37. The method of claim 31 , wherein the compound is in a tablet or capsule that is administered orally at least four times daily.

38. The method of claim 31, wherein the compound is administered to the subject four times daily for not more than 4 days.

39. The method of claim 31, wherein the compound is administered in a pulse preceded by a water fast.

40. The method of claim 39, wherein the water fast is about 3 days long.

41. The method of claim 40, wherein the pulse is between about 1 and 2 days.

42. The method of claim 41, wherein the pulse comprises administering a dose multiple times per day.

43. The method of claim 42, wherein the dose is administered once every hour during the pulse.

44. The method of claim 43, wherein the dose is administered in enteric capsules.

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