Sagerinic acid and compositions containing same, for use in the prevention and / or treatment of senescence-related disorders in a human or animal

Abstract

The invention relates to sagerinic acid and the compositions containing it, for use in the prevention and / or treatment of disorders related to senescence in humans and animals.

Description

[0001]SAGERINIC ACID AND COMPOSITIONS CONTAINING IT FOR USE IN THE PREVENTION AND / OR TREATMENT OF SENESCENCE-RELATED DISORDERS IN A HUMAN OR ANIMAL The invention relates to sagerinic acid and compositions containing it for use in the prevention and / or treatment of senescence-related disorders in humans and animals. Aging is a multifactorial process characterized by the progressive loss of physiological functions, leading to an increase in vulnerability to age-related diseases. Several theories have been proposed to explain the nature of aging and there is now a consensus on how to categorize the different mechanisms involved in aging. One of the best known identifies free radicals produced by mitochondrial metabolism as the cause of cellular disorders and damage to genetic material (1).Another well-known axis of aging concerns the alteration of genetic material over time. There is several evidence supporting that epigenetic modifications, such as DNA methylation, non-coding RNA and histone modifications play a key role in the molecular mechanism of aging. Caloric restriction is the intervention that, in all animal species on which it has been applied, has produced beneficial effects on the health of aging organisms and significantly increased their longevity. It has been shown that sirtuins, which are histone deacylation enzymes that protect genetic material from degradation induced by various factors, are strongly positively involved in the effects of caloric restriction (2). Another axis of aging concerns protein glycation (3).Generally speaking, proteins all interact with reducing sugars during a passive, spontaneous reaction, not involving an enzyme, to give condensed compounds, in a more or less irreversible way. This reaction is the Maillard reaction, the mechanisms of which were first described by JE Hodge in 1953 (4). This reaction gives rise to multiple rearranged compounds (Amadori bodies among others). When this reaction is carried to completion, we then speak of "Advanced Glycation Endproducts" or AGEs. This reaction is universal. It occurs in all cases, but to a greater or lesser extent depending on the sugars involved and also the number of free amino radicals present on the protein molecule. This reaction is amplified by heat and depends on the concentration of the sugars involved present in the medium.During glycation, proteins undergo "crosslinks" that alter their functionality and stiffen structural proteins such as collagen and elastin. These proteins are normally eliminated by proteases that allow their renewal. The proteins are less sensitive to the effect of proteases the more advanced the glycation. Slow-renewing proteins, such as those in joints or the brain, therefore undergo particularly strong glycation linked to this slow renewal. Aging therefore passively leads to increased glycation of slow-renewing proteins. Diabetes leads to an identical result due to high blood sugar. In both cases, there is a weakening of organs whose functions depend on connective tissue, particularly the vascular system.Thus, diabetic disease is generally associated with various angiopathies, particularly at the ocular and renal level. Disorders associated with glycation, whether normal, i.e. correlated with physiological aging, or pathological as is the case in diabetes (5), are very varied and of varying severity. We can cite the loss of elasticity of blood and lymphatic vessels which leads to all forms of angiopathies associated with diseases such as strokes, atheroma, macular degeneration, renal failure (6), respiratory failure. We can also cite aesthetic integumentary damage such as sagging of the skin. Overall, it also seems established that the glycation of collagen, the most abundant protein in mammals, would be a reliable marker of biological age and would make it possible to determine life expectancy (7, 8).Aging induces remodeling of large arteries. These age-related alterations may be associated with increased activity of metalloproteinases, particularly collagenases and elastases, in large-diameter arteries (9). It is therefore important that the activity of these enzymes is perfectly regulated. This regulation may be provided by the protection of arterial connective tissue proteins, mainly collagen and elastin, against the excessive action of these enzymes.As a consequence of the above, to act effectively against the aging of the organism, it would be desirable to act simultaneously on what appear to be the four fundamental axes of aging: - Oxidative stress by the use of anti-radical molecules - Glycation of proteins by molecules capable of reversing the glycation already installed - Protection of DNA by activation of sirtuins - Protection of connective tissue by inhibition of proteases which degrade it and in particular elastases and collagenases. Molecules capable of acting simultaneously on one or more of these four axes have already been described. Antioxidants and free radical traps Many antioxidants and free radical traps have been described since the beginning of the twentieth century, particularly in the plant kingdom in which polyphenols are the most frequent representatives.There are also molecules such as water-soluble vitamin C and fat-soluble vitamin E, which have good anti-radical power. A simple and rapid method for evaluating the anti-radical effect uses a stable free radical, 2,2-diphenyl 1-pyrrylhydrazyl (DPPH), whose degradation by the molecule to be tested is measured by colorimetry. Glycation inhibitors and protein deglycants A large number of molecules are now known to inhibit protein glycation (10). Most act simply by antioxidant effect, since the glycation reaction includes an oxidative phase. Thus, for example, a large number of polyphenols have been recognized as being able to be used as inhibitors of the protein glycation reaction.Others, such as aminoguanidine, carnosine, a monopeptide from red meat, and pyridoxamine, act as competitive inhibitors through their amino radical, with respect to the amino radical of the oses involved in glycation. Protein glycation inhibitors are of interest for health by limiting this reaction and producing fewer new cross-links. However, glycation causes a slowdown in protein turnover, inducing their accumulation and the functional degradation of the organs to which they are linked. Some proteins are slowly renewed, notably connective tissue proteins, collagen and elastin, which play a leading role in the viscoelastic properties of the cardiovascular system, joints and the dermis.It is therefore of great importance to discover molecules capable of achieving this reversion of glycation, in order to cut the pre-existing cross-links and allow tissues to regain, at least in part, their original viscoelastic properties. In particular, the decrease in elasticity of the cardiovascular system is the most important problem during aging and the main cause of this decrease is the formation of AGEs and cross-links between protein molecules. The use of a glycation inhibitor does not allow this system to regain the elasticity it has lost over time. For this, an agent capable of cutting cross-links is necessary. There is therefore a clear need for products capable of breaking the protein cross-links created by glycation and having no harmful side effects. The applications of such products are considerable.Among the most important, we can cite the fight against angiopathies and in particular the nephropathies, cardiopathies and retinopathies attached to them, the improvement of the blood and lymphatic vascular functioning of the elderly subject, the improvement of the elasticity of the skin leading to an improvement in appearance, the fight against the loss of elasticity of tendons and cartilage, leading to reductions in mobility, due to inflammation and pain in the joints. All these pathologies are associated with age in normal subjects and are aggravated by diabetes. We can also cite the improvement of venous and lymphatic functioning during the resolution of trauma by improving the functionality of connective tissue proteins thanks to the reduction of their glycation rate.Finally, aging is accompanied by a degradation of vascular connective tissue leading to stiffening of blood and lymphatic vessels and the creation of atheromatous plaques. Certain molecules of the thiazolium group have been described in US Patent 5853703A (22, 23) as possessing this activity in vitro and in animal pharmacology studies. Patent WO2011042890A2 discloses a family of polyphenolic molecules possessing a protein deglycation effect (11). Sirtuin activators (12) NAD+-dependent protein lysine deacylases of the sirtuin family regulate various physiological functions, from energy metabolism to stress responses.Human Sirtuin isoforms, SIRT1-7, which are particularly activated during caloric restriction are considered attractive therapeutic targets for aging-related diseases, such as type 2 diabetes, inflammatory and neurodegenerative diseases. Pharmacological activators of sirtuin 1 and other isoforms have been described, and initial clinical trials have been conducted. Connective tissue protectants (13) Epidemiological studies have suggested an association between the consumption of foods or beverages containing polyphenols and the prevention of certain human diseases, such as chronic obstructive pulmonary and cardiac diseases, chronic inflammation, as well as the reduction of the risk of many types of cancer.It now appears to be proven that polyphenols protect extracellular matrix proteins against attacks from overexpressed proteases during exposure to free radicals and during inflammation. However, none of these molecules alone can act simultaneously on the four fundamental axes of aging, which are: - Oxidative stress through the use of anti-radical molecules - Glycation of proteins by molecules capable of reversing the glycation already established - Protection of DNA by activation of sirtuins - Protection of connective tissue by inhibition of proteases that degrade it, particularly elastases and collagenases. Furthermore, sagerinic acid is a molecule that was discovered and described for the first time in common sage (Salvia officinalis) by Yinrong Lu and L. Yeap Foo in 1999 (17).Sagerinic acid has since been identified in a large number of plant species of the Lamiaceae family such as Melissa officinalis (18), Rosmarinus officinalis (19), Orthosiphon grandiflorus (20), Helicteres hirsuta (21), Plectranthus amboinicus (15). Sage and lemon balm are plants with multiple applications and properties and generally, these multiple effects are due to the presence in the extracts of different molecules. The efficacy of sagerinic acid as an inhibitor of COVID19 was predicted by an in silico simulation, but no material evidence of this activity has been produced (14). Furthermore, US patent US 8,105,636 B2 (15) claims a composition comprising an extract of Plectranthus amboinicus as an anti-inflammatory in which sagerinic acid is cited as one of the possible active ingredients. The potential antileishmanial and anti-inflammatory activity of sagerinic acid has been described (16).The invention is based on the fortuitous discovery of the effects of sagerinic acid on these four fundamental axes of aging: during in vitro tests, it was observed that, surprisingly, a single molecule, sagerinic acid, possesses the four properties described above. The invention applies to the prevention and treatment of disorders linked to senescence.Among these, we can cite, without this list being exhaustive: - connective tissue diseases affecting cartilage such as articular osteoarthritis and rheumatoid arthritis in their non-inflammatory components (degradation of connective tissue and not immune reactions leading to inflammation), - vascular connective tissue diseases: strokes, vascular nephritis and retinitis, macular degeneration, atheromatous disease, - diseases directly linked to protein glycation such as cataracts, Alzheimer's disease, skin healing disorders, presbycusis, - aesthetic skin disorders such as lipofuscin accumulation, pigment spots, erythema, loss of skin elasticity.Sagerinic acid may be applied in pure form or in extract form and combined with excipients suitable for its use: oral, parenteral, topical, rectal or inhalation. Thus, the invention provides sagerinic acid for use in the prevention and / or treatment of disorders related to senescence in a human or animal being. In a first embodiment, these disorders are due to oxidative stress. In a second embodiment, these disorders are due to protein glycation. In a third embodiment, these disorders are due to DNA damage. In a fourth embodiment, these disorders are due to arterial connective tissue protein damage. The invention also provides sagerinic acid for use as an antioxidant and / or free radical scavenger. The invention also provides sagerinic acid for use as a protein deglycating agent.The invention further provides sagerinic acid for use as a DNA protection agent. But also, the invention provides sagerinic acid for use as a protease inhibition agent. In a preferred embodiment, the invention provides sagerinic acid for use in the simultaneous prevention and / or treatment of disorders due to oxidative stress, and disorders due to protein glycation, and disorders due to DNA damage and disorders due to arterial connective tissue protein damage, in a human or animal. Another subject of the invention is a composition comprising sagerinic acid for use in the prevention and / or treatment of senescence-related disorders in a human or animal. In a first embodiment of the composition of the invention, the disorders are due to oxidative stress.In a second embodiment of the composition of the invention, the disorders are due to protein glycation. In a third embodiment of the composition of the invention, the disorders are due to DNA damage. In a fourth embodiment of the composition of the invention, the disorders are due to arterial connective tissue protein damage. A preferred subject of the invention is a composition comprising sagerinic acid for use in the simultaneous prevention and / or treatment of disorders due to oxidative stress, and disorders due to protein glycation, and disorders due to DNA damage and disorders due to arterial connective tissue protein damage, in a human or animal being. The invention will be better understood and the characteristics and advantages thereof will appear more clearly upon reading the explanatory description and the implementation examples which follow.Examples Example 1: Preparation of purified sagerinic acid 170 g of dried lemon balm leaves (Melissa officinalis) are macerated in 2 L of deionized water at room temperature and with constant stirring for 24 hours. The extractive solution is separated by filtration on paper, then lyophilized. The weight of the extraction residue is 32.3 g, i.e. a yield of 19%. 10 g of lyophilized extract are dissolved in 50 mL of deionized water and this solution is chromatographed on a column 45 mm in diameter and 24 cm long containing approximately 200 g of Daïaon HP20 resin (Thermo Scientific ref.046488.A1) with 2 L of deionized water as eluent. The 2 L eluted from the column are discarded. Then eluted with 2 L of methanol (Thermo Scientific ref. L13255). All of the eluate is collected and evaporated under reduced pressure until the methanol has completely evaporated.The remaining aqueous phase is made up to 100 mL with deionized water, 10 mL of concentrated hydrochloric acid (Honeywell Fluka, ref. 320331) is added and countercurrently extracted with 3 times 100 mL of ethyl acetate (Roth Ref. 73361). The organic phase is dried over anhydrous sodium sulfate (Sigma Aldrich, ref. 238597), filtered and evaporated to dryness. Approximately 700 mg of partially purified sagerinic acid is obtained. The 700 mg of crude sagerinic acid are then chromatographed in batches of 100 mg dissolved in 1 mL of deionized water by preparative HPLC on a C18 column of 22 mm diameter and 250 mm length (Vydac Denali 10µm) by the following gradient: Flow rate: 20 mL / min Solvent A: Deionized water containing 0.1% formic acid (Fischer Chemical A117-50) Solvent B: methanol (Thermo Scientific ref. L13255). 10 ml fractions are collected. The fraction containing sagerinic acid is identified by its UV spectrum (max). It is retained and the other fractions are discarded. After passing the 700 mg, the fractions are combined and evaporated to dryness under reduced pressure. A residue of 165 mg is obtained. The remaining 165 mg are then chromatographed in the same way as before. 23 mg of 92% pure sagerinic acid is finally obtained. Example 2: Preparation of lemon balm extract enriched with sagerinic acid 170 g of dry lemon balm leaves are macerated in 2 L of deionized water at room temperature and with constant stirring for 24 hours. The extractive solution is separated by filtration on paper, then lyophilized. The weight of the extraction residue is 32.3 g, i.e. a yield of 19%.10 g of lyophilized extract are dissolved in 50 mL of deionized water and chromatographed on a column 45 mm in diameter and 24 cm long containing approximately 200 g of Daïaon HP20 resin (Thermo Scientific ref. 046488.A1) with 2 L of deionized water as eluent. The 2 L eluted from the column are discarded. Elution is then carried out with 2 L of methanol (Thermo Scientific ref. L13255). All of the eluate is collected and evaporated under reduced pressure until the methanol has completely evaporated. The remaining aqueous phase is made up to 100 mL with deionized water, 10 mL of concentrated hydrochloric acid (Honeywell Fluka, ref. 320331) is added and countercurrently extracted with 3 times 100 mL of ethyl acetate (Roth Ref. 73361). The organic phase is dried over anhydrous sodium sulfate (Sigma Aldrich, ref.238597), filtered and evaporated to dryness. Approximately 700 mg of lemon balm extract containing 8% sagerinic acid is obtained.Example 3: Evaluation of the effects of sagerinic acid and ascorbic acid on oxidative stress 3.1. Principle The principle of the method for evaluating the effect of sagerinic acid compared to ascorbic acid as a reference molecule is based on the elimination of a stable free radical, 2,2-Diphenyl-1-picrylhydrazyl (DPPH Sigma Aldrich ref. D9132). 3.2. Apparatus and materials - Jasco V-730 UV-Visible spectrophotometer with double beam - Stopwatch - Standard laboratory equipment. - Precision balance: ML104 (Mettler Toledo). - Automatic pipettes with variable volume. 3.3. Reagents - Monopotassium phosphate (Riedel de Haen, ref. 04243) - Dibasic sodium phosphate dihydrate (Sigma Aldrich ref.30435) - Purified water - DPPH (Sigma Aldrich, ref. D9132) - Methanol Optima® LC / MS Grade (Fisher Chemical, ref.A456-212) - Toluene, Reference (Carl Roth ref.4445.1) - Ethyl alcohol Pharmethyl 96°, (Cristalco ref. 11371014) 3.4.Preparation of reagent solutions: - Phosphate buffer pH 7 Dissolve 0.177g of monopotassium phosphate and 0.730g of dibasic sodium phosphate dihydrate in 50ml of purified water. Adjust the pH if necessary. - 2.2-diphenyl 1-pyrrylhydrazyl (DPPH) solution in methanol Prepare a solution of exactly 0.0027% (w / V) DPPH in methanol. - Antioxidant solution Prepare a solution of exactly 0.1% (w / V) ascorbic acid (Riedel-de-Haën, ref.33034) in ethyl alcohol. Dilute this solution to 0.0007% (V / V) in ethyl alcohol. A solution of exactly 0.1% (w / v) sagerinic acid (prepared according to Example 5.1) is prepared in water. This solution is diluted to 0.0007% (v / v) in water. 3.5. Procedure 3.5.1. Control Solution: In a series of hermetically sealed pill boxes, the following are introduced: 3.5.2. Test: In a series of hermetically sealed pill boxes, the following are introduced: For the sample and the blank: - After addition, stir for 5 min on a magnetic stirrer - Add 4 ml of toluene - Stir for 30 seconds on a magnetic stirrer - Leave to settle for 20 seconds to 1 minute (if necessary) - Collect the organic phase on top using an automatic pipette, taking care not to collect any aqueous phase - Read the OD of the organic phase on a spectrophotometer at 519 nm 3.6. Results It can be seen from these results that sagerinic acid has an effect almost twice that of ascorbic acid on the elimination of free radicals and therefore on the prevention and / or treatment of disorders due to oxidative stress. Example 4: Evaluation of the effects of sagerinic acid and Alagebrium as a protein deglycant. 4.1. Principle The test products in 0.25% (w / V) solution are left in contact with a 0.375% (w / V) solution of 1-phenyl-1,2-propanedione (PPD) for 24 hours at 37°C. An HPLC assay, at 229 nm after passage on an Ascentis column ®Express C18, is carried out at T0 and then after 24 hours to determine the benzoic acid content. Calculating the percentage of µmole of benzoic acid / µmole of PPD in the reaction medium determines the activity of the deglycants. 4.2. Apparatus and materials - Ultimate 3000 HPLC system (Thermo Scientific - Dionex) with diode array detector and column oven. - Ascentis HPLC column ®Express C18 (Supelco Ref 53816-U) Dimensions: 15 cm x 3.0 mm Particle size: 2.7 µm - BD 53 / E2 oven (Binder). - PVDF (Polyvinyl fluoride) filters with a porosity of 0.45 µm (Millex®-HV Ref SLHVX13NL). - Standard laboratory equipment. - Precision balance: ML104 (Mettler Toledo). - Automatic pipettes with variable volume. 4.3. Reagents - Optima® LC / MS Water Suitable for UHPLC-UV (Fisher, ref.W6-212) - Optima® LC / MS Grade Acetonitrile, (Fisher Chemica, ref.A955-212) - Optima® LC / MS Grade Methanol, (Fisher Chemical, ref. A456-212) - Pharmethyl 96° Ethyl Alcohol (Cristalco, ref.11371014) - Sodium Acetate Trihydrate (Sigma Aldrich ref. S8625) - Acetic Acid (Carl Roth ref. HN55.1) - Benzoic Acid, Reference 242381 (Sigma Aldrich ref.242381) - Monopotassium Phosphate (Riedel de Haen, ref.04243) - Dibasic Sodium Phosphate dihydrate (Sigma Aldrich ref. 30435) - 1-Phenyl-1,2-Propanedione (PPD) (Aldrich ref.223034) 4.4.Preparation of reagent solutions - Phosphate buffer pH 7.4 Dissolve 0.178 g of monopotassium phosphate and 0.955 g of dibasic sodium phosphate dihydrate in 100 ml of purified water. Adjust the pH if necessary. - Phosphate buffer pH 7.4 - Methanol Prepare a mixture with 50 ml of phosphate buffer pH 7.4 and 50 ml of methanol. - 1-Phenyl-1,2-Propanedione (PPD) solution Prepare a solution of exactly 0.375% (w / V) PPD in the phosphate buffer pH 7.4 - methanol mixture (50 / 50). 4.5. Preparation of a test product solution Prepare a solution of approximately exactly 0.25% (w / V) of the test product in the phosphate buffer pH 7.4 - methanol mixture (50 / 50). Test product: Alagebrium chloride (MedChem Express, ref. HY-106024B), Sagerinic acid (prepared according to example 5.1.). 4.6. Benzoic acid calibration range 4.6.1. Standard solutions A stock solution of benzoic acid is prepared at approximately exactly 0.08% (w / V) in ethyl alcohol.This solution is diluted 1 / 10 (V / V) in purified water (D). Then (D) is diluted 1 / 5, 2 / 5, 3 / 5 and 4 / 5 (V / V) in purified water. (D1, D2, D3 and D4). D1, D2, D3, D4 and D are passed through PVDF filters with a porosity of 0.45 µm. 4.6.2 Mobile phases A: Sodium acetate buffer: 3.18 g of sodium acetate trihydrate are weighed, 10 ml of acetic acid are added and the volume is made up to 500 ml with “Optima” water, and the volume is stirred until completely dissolved. B: Acetonitrile 4.6.3 Analytical conditions Column: Ascentis. ® Express C1815 cm x 3.0 mm 2.7 µm Flow rate: 0.5 ml / min at 25°C Mode: gradient (see table 1) Detection: ultraviolet (UV) at 229 nm Injections: 10 µl for solutions D1, D2, D3, D4 and D. Table 1: A linear calibration line is established: area of ​​the benzoic acid peak, having a retention time of 9.17 min and observed at 229 nm, as a function of the concentration in % (w / V). 4.7. Action on PPD 4.7.1. Procedure: In a series of hermetically sealed pill boxes, the following are introduced: Shake after each addition. Take 0.05 ml from each pillbox to measure the benzoic acid content at T0. The pillboxes are closed and placed in an oven at 37°C for a minimum of 21 to a maximum of 24 hours. The same volume will be taken for determination upon removal from the oven. It is necessary that the time at 37°C be exactly the same for the control and all the tests, so they are prepared as they are carried out. The HPLC determinations of benzoic acid are also carried out as they are carried out. 5.4.7.2. HPLC determination of released benzoic acid: Dilute the 0.05 ml samples 1 / 5 (V / V) in purified water, i.e. by adding 0.2 ml. They are passed through PVDF filters with a porosity of 0.45 µm before injection. 5.4.7.2.1. Analytical conditions: Column: Ascentis ® Express C1815 cm x 3.0 mm 2.7 µm Flow rate: 0.5 ml / min at 25°C Mode: gradient (see table 2) Detection: ultraviolet (UV) at 229 nm Injections: 10 µl Table 2: 4.8 Results Using the calibration line, we calculate the benzoic acid content in g / 100 ml of reaction medium for - PPD control at T0 and 24 hours: T T0 and T 24h - test at T0 and 24 hours: ET0 and E24h These concentrations are transformed into µmol of benzoic acid / ml of reaction medium (PM benzoic acid = 122.12) TT0 x 10000 / 122.12 The PPD concentration in µmol / ml of reaction medium is as follows: 0.375 x 1000000 / 100 x 2 x 148.16 = 12.655 (PM PPD = 148.16) The % of µmol of benzoic acid / µmol of PPD in the reaction medium is calculated: TT0 x 10000 x 100 / 122.12 x 12.655 The values ​​obtained for the control and the test at the different times are compared. The deglycating activity of sagerinic acid was compared to that of Alagebrium as a reference molecule. It is seen from these results that sagerinic acid has good protein deglycating activity, in addition to its excellent free radical scavenging activity. Example 5: Evaluation of the effects of sagerinic acid and resveratrol on sirtuin 1 5.1. Principle The SIRT1 Inhibitor / Activator Screening Kit deacetylates the substrate with SIRT1, followed by cleavage of the deacetylated substrate to release the fluorescent group, which is detected by fluorimetry at Ex / Em = 400 / 505 nm. The SIRT1 activator enhances SIRT1 activity resulting in a higher fluorescent signal compared to the control. This kit provides a rapid, simple, sensitive and reliable assay suitable for high-throughput screening of SIRT1 activators. 5.2. Apparatus and materials - Gemini EM spectrofluorimeter - Stopwatch - Common laboratory equipment. - Precision balance: ML104 (Mettler Toledo). - Automatic pipettes with variable volume - Innovens 28EU1 Jouan / Thermo5 oven.3 Reagents - Kit solutions: Reference ab283377 (Abcam) ^ Buffer ^ Substrate ^ SIRT1 ^ NAD ^ DTT - DMSO Reference 34869 (Honeywell Riedel-de-Haën) 5.4. Preparation of reagent solutions SIRT1 enzyme: Store at -80°C. NAD: Store at -80°C. Avoid repeated freezing / thawing. Mix 2 μL of NAD stock solution with 58 μL of DTT-free buffer. 1 M DTT: Store at -20°C. Thaw and keep on ice during use. Buffer: Store at 4°C or -20°C. Warm to 37°C. Must be prepared just before use. Preparation of DTT buffer solution: Mix 0.002 ml of DTT with 0.998 ml of buffer. Developer: Store at -20°C. Avoid repeated freezing / thawing. Keep on ice during use. 5.5. Preparation of a solution of the product to be tested (activator) A solution of exactly 2.28% (w / V) of activator (resveratrol (Sigma Aldrich, ref.R5010) and sagerinic acid prepared according to example 5.1.) in DMSO is prepared. Dilute to 1 / 250 èmein the DTT buffer solution. 5.6. Procedure Preparation of the enzymatic solution: In 3 separate wells, make the following additions: SIRT1 enzyme 2 μL Screening Compounds, Inhibitor Control, Enzyme Control and Blank Control Preparations: 1. Into the SIRT1 Enzyme Solution wells, add: - 25 μl of the undiluted Activator Solution (2.28% solution) in one well as Activator Control (AC) - 25 μl of DTT-free Buffer in one well as Enzyme Control (EC) - 25 μl of the diluted Activator Solution as Test (S) 2. In another well, add 50 μl of DTT-free Buffer as a blank control (without enzyme). 3. Mix well and incubate the plate for 5 min. at 37°C Substrate Preparation: After incubation, make the following additions to each of the wells: Mix and incubate at 37°C for 30-60 min. Developing: Add 10 μl of developer to each well. Mix well and incubate for 10 min. at 37°C, protected from light. Measurement: Read the fluorescence (Ex / Em = 400 / 505 nm). 5.7. Results The activation of sirtuin 1 by sagerinic acid was compared to that of resveratrol as a reference molecule. It can be seen from these results that sagerinic acid has good activity on sirtuin activation, i.e. a good effect in preventing and / or treating DNA damage, in addition to good protein deglycating activity, and excellent free radical scavenging activity. Example 6: Evaluation of the effects of sagerinic acid and hamamelitannin as connective tissue protein protectors. 6.1. Principle A collagenase activity assay kit (Sigma Aldrich MAK293) is used. The kit allows screening of collagenase inhibitors by measuring collagenase activity using a synthetic peptide (FALGPA) that mimics the structure of collagen. 6.2. Apparatus and materials - Jasco V-730 UV-Visible spectrophotometer with double beam - Stopwatch - Standard laboratory equipment. - Precision balance: ML104 (Mettler Toledo). - Automatic pipettes with variable volume - Innovens 28EU1 Jouan / Thermo 6.3 oven.Reagents - Kit solutions: Reference MAK293-1KT (Sigma Aldrich) ^ Collagenase buffer (MAK293A-KC) ^ Collagenase solution (MAK293B-KC) ^ Collagenase substrate (MAK293C-KC) - Ethyl alcohol Pharmethyl 96°, Reference 11371014 (Cristalco) - Purified water 6.4. Preparation of a solution of product to be tested (inhibitor) A solution of exactly 1% (w / V) of hamamelitannin (Extrasynthèse ref. 0958) is prepared in ethyl alcohol. A solution of exactly 1% (w / V) of sagerinic acid (prepared according to example 5.1.) is prepared in purified water. 6.5. Procedure 6.5.1. Control Solution In a series of hermetically sealed pill boxes, we introduce: - After addition, shake quickly by hand - Place in an oven at 37°C for 30 minutes - Add 1.6 mL of purified water. Mix. - Read the OD: Sample against Blank, using a spectrophotometer at 345 nm. 6.5.2. Results Activity on collagenases These results show that sagerinic acid has an inhibitory effect on proteases, represented here by collagenases, more than twice that of the reference molecule, hamamelitannin. This is in addition to excellent free radical scavenging activity, good activity on sirtuin activation, i.e. a good effect in preventing and / or treating DNA damage, and good protein deglycating activity.Sagerinic acid therefore has excellent activity, on its own, in preventing and / or treating disorders related to senescence in humans or animals because it acts simultaneously on the four fundamental axes of aging, namely oxidative stress thanks to its excellent anti-radical action, protein glycation thanks to its good capacity to reverse the already established glycation of proteins, DNA protection through its good capacity to activate sirtuins and connective tissue protection thanks to its excellent activity of inhibiting proteases that degrade it, particularly elastases and collagenases. The application of sagerinic acid to the treatment of the four main axes of aging will be done according to doses defined with regard to what would be necessary to administer the four reference molecules used together.The recommended daily oral dose of ascorbic acid is between 200 mg and 1 g per day (24), that of resveratrol is between 250 mg and 1 g per day (25), that of hamamelitannin is between 700 mg and 2 g per day (26 and 27) and that of Alagebrium is between 100 mg and 300 mg per day (28). Thus, a daily oral dose of sagerinic acid of between 10 mg and 2 g per day provides the recommended amounts of ascorbic acid and resveratrol and of hamamelitannin and Alagebrium. An advantage of the invention is therefore that a single dose of sagerinic acid corresponds to separate doses of ascorbic acid, resveratrol, Alagebrium and hammamelitannin. Example 7. Preparation of capsules of Melissa officinalis extract enriched with sagerinic acid. 1 kg of dry Melissa officinalis leaf extract obtained according to Example 2 is thoroughly mixed with 10 g of sodium carbonate (E500) in a knife mill.This mixture is then used to fill capsules No. 00 (0.9 mL). The capsule thus contains 550 mg of extract or 44 mg of sagerinic acid. Example 8. Galenic formula of a gel intended to treat aesthetic disorders, suitable for use on the body and face. Purified water: 91.69% Carbomer (Carbopol 980, Lubrizol): 2% Extract of dried leaves of Melissa officinalis as described in example 1: 5% Benzyl alcohol (Geogard 221, Lonza): 0.87% Dehydroacetic acid (Geogard 221, Lonza): 0.09% Sodium hydroxide (Sigma-Aldrich): 0.35% Bibliographic references 1. Wickens AP. Ageing and the free radical theory. Respir Physiol., 2001, 128(3), 379-91. 2. Grabowska W, Sikora E, Bielak-Zmijewska A. Sirtuins, a promising target in slowing down the aging process. Biogerontology. 2017, 18(4), 447-476. 4. John E. Hodge Chemistry of Browning Reactions in Model Systems. Agricultural and food chemistry, 1953, 1 (15), 928-943. 5. Wautier J.L., Guillausseau P.J. Advanced Glycation end products, their receptors and diabetic angiopathy. Diabetes Metab (Paris), 2001, 27, 535-542. 6. Sakata N., Noma A., Yamamoto Y., Okamoto K., Meng J., Takebayashi S., Nagai R. and Horiuchi S.Modification of elastin by pentosidine is associated with the calcification of aortic media in patients with end-stage renal disease. Nephrol. Dial. Transplant.2003, 18, 1601-1609 7. Pageon H., Asselineau D. An in vitro approach to the chronological aging of skin by glycation of the collagen. Ann. N.Y. Acad. Sci.2005, 1043, 529-532 8. Sell D.R. et al. Longitudinal determination of skin collagen glycation and glycoxydation rates predicts early death in C57BL / 6NNIA mice. FASEB Journal, 2000, 14, 145-156 9. 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Claims

CLAIMS 1. Sagerinic acid for use in the prevention and / or treatment of senescence-related disorders in a human or animal.

2. Sagerinic acid according to claim 1 characterized in that the disorder is due to oxidative stress.

3. Sagerinic acid according to claim 1 characterized in that the disorder is due to protein glycation.

4. Sagerinic acid according to claim 1 characterized in that the disorder is due to DNA damage.

5. Sagerinic acid according to claim 1 characterized in that the disorder is due to arterial connective tissue protein damage.

6. Sagerinic acid for use as an antioxidant and / or free radical scavenger.

7. Sagerinic acid for use as a protein deglycating agent.

8. Sagerinic acid for use as a DNA protection agent. 9.Sagerinic acid for use as a protease inhibiting agent.

10. Sagerinic acid for use in the prevention and / or treatment, simultaneously, of disorders due to oxidative stress, and disorders due to protein glycation, and disorders due to DNA damage and disorders due to arterial connective tissue protein damage, in a human or animal.

11. Composition comprising sagerinic acid for use in the prevention and / or treatment of senescence-related disorders in a human or animal.

12. Composition according to claim 10 characterized in that the disorder is due to oxidative stress.

13. Composition according to claim 10 characterized in that the disorder is due to protein glycation.

14. Composition according to claim 10 characterized in that the disorder is due to DNA damage. 15.Composition according to claim 10 characterized in that the disorder is due to an alteration of the proteins of the arterial connective tissue.

16. Composition comprising sagerinic acid for use in the prevention and / or simultaneous treatment of disorders due to oxidative stress, and disorders due to protein glycation, and disorders due to DNA damage and disorders due to an alteration of the proteins of the arterial connective tissue, in a human or animal being.

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