Iridoid compound as well as preparation method and anti-aging application thereof

Abstract

The invention discloses an anti-aging compound which is a new compound extracted from fructus forsythiae and is named as cyclic pentacarbazide acid ester. The invention also discloses a preparation method of the forsythia suspensa extract, which comprises the following steps: extraction: carrying out reflux extraction on a dried medicinal material by using an ethanol water solution, filtering, and recovering under reduced pressure to remove a solvent, so as to obtain the forsythia suspensa extract; according to the present invention, the pure product of the cyclocardioside acid ester is obtained through the steps of macroporous resin separation, C18 separation, gel purification and target fraction collection, and the experiment results of Caenorhabditis elegans show that the cyclocardioside acid ester can prolong the life cycle of the Caenorhabditis elegans, and the cyclocardioside acid ester and the Caenorhabditis elegans have significant anti-aging effects.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, specifically a method for preparing cyclopentafloridoside acid ester from Chinese medicinal materials, and the application of cyclopentafloridoside acid ester as an anti-aging drug. Background Art

[0002] With the intensification of global population aging, the demand for delaying the aging process is increasing day by day. Aging is a complex biological process involving the interaction of multiple factors such as genetics, environment, and lifestyle. Although aging itself is not a disease, it is closely related to the increased incidence of various chronic diseases, such as cardiovascular diseases, diabetes, neurodegenerative diseases, etc. Therefore, exploring effective anti-aging strategies can not only improve the quality of life of individuals but also have a profound impact on public health. Scientific research shows that antioxidants, free radical scavengers, and certain natural compounds can delay cell damage by reducing oxidative stress, thus playing a positive role in the anti-aging process. In addition, by regulating cell metabolic pathways, promoting DNA repair mechanisms, or enhancing autophagy function, some compounds can effectively slow down aging-related changes. These findings have promoted the development of the anti-aging drug research and development field, prompting scientists to search for safer and more effective anti-aging ingredients. It is worth noting that modern medical research increasingly focuses on how to find answers from nature, especially those active ingredients derived from plants. Compounds from plant sources have become a new hot spot in anti-aging research due to their rich biological activities and low toxicity. They not only perform well in the laboratory environment but also show good tolerance and acceptance in practical applications. For example, polyphenols (such as resveratrol), flavonoids (such as quercetin), and terpenoids (such as ursolic acid) have all been widely studied and shown significant antioxidant, anti-inflammatory, and cell damage protection abilities. Such compounds usually have low toxicity and high bioavailability, making them important candidates for developing new anti-aging products.

[0003] In recent years, compounds derived from plants have become a hot topic in anti-aging research due to their rich bioactivities and low toxicity. Many studies have shown that polyphenols (such as resveratrol), flavonoids (such as quercetin), and terpenoids (such as ursolic acid) have significant antioxidant, anti-inflammatory, and cell damage protection abilities. For example, resveratrol, a polyphenol naturally present in grape skins, has been proven to activate the SIRT1 protein, an enzyme closely related to lifespan extension. Another study pointed out that quercetin can reduce inflammatory responses by inhibiting the NF-κB signaling pathway, thereby playing a positive role in anti-aging. At the same time, NMN (nicotinamide mononucleotide), a derivative of vitamin B3, has shown excellent performance in increasing NAD+ levels and has obtained dual certifications of FDA-GMP and ISO22000. Clinical verification shows that the level of inflammatory factor IL-6 in long-term users is reduced by 40%, which further proves its potential anti-aging effect. In addition to the specific cases mentioned above, anti-aging compounds derived from plants have other advantages. First, since these compounds come from nature, they often have lower toxicity than synthetic chemical drugs and less side effects on the human body. Second, compounds derived from plants are more easily absorbed and utilized by the human body because they usually exist in natural forms, with complex structures but easy to recognize and process. Moreover, since plants have been used as food and medicine in human history for a long time, the public has a relatively high acceptance of plant-derived products, which is a huge advantage for promoting new anti-aging products. In summary, compounds derived from plants are becoming an important resource for developing new anti-aging products due to their unique bioactivities, safety, and good acceptability.

[0004] In the preliminary screening process of researching anti-aging compounds derived from plants, the inventors found that the extract of Forsythia suspensa showed considerable anti-aging characteristics, which is not the same as its traditional medicinal uses. In traditional medicine, Forsythia suspensa (the dried fruit of Forsythia suspense (Thunb.) Vahl, a plant of the Oleaceae family) is divided into Qingqiao and Laoqiao according to the harvesting time. Forsythia suspensa has the effects of clearing heat and detoxifying, dissipating swelling and nodules, and dispersing wind-heat, and can be used for carbuncles, scrofula, breast abscess, erysipelas, wind-heat colds, the initial stage of warm diseases, the entry of warm heat into the nutrient system, high fever and restlessness, coma and maculae, and stranguria with pain, but it has no anti-aging uses. It is speculated that there may be a class of compounds with non-dominant contents whose functions are magnified and manifested in the modern drug screening system. For the purpose of finding more anti-aging compounds derived from plants, the inventors further studied the chemical constituents of Forsythia suspensa and finally found a previously unreported compound, which belongs to the type of iridoid trimer and has obvious anti-aging effects. Summary of the Invention

[0005] The inventors of the present invention discovered a iridoid trimer structure from Forsythia suspensa Vahl. Specifically, the present invention provides an iridoid compound represented by Formula 1 or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof.

[0006]

[0007] In the formula, X independently represents O, S or CH2, and R independently represents H, Glc, Fru, Gal or Xyl.

[0008] In one embodiment of the present invention, X represents O, and R represents H or Glc.

[0009] In one embodiment of the present invention, the compound represented by Formula 1 has the following structure.

[0010]

[0011] Another aspect of the present invention provides the use of the above compound in the preparation of a drug for anti-aging.

[0012] The present invention also provides a pharmaceutical composition, which comprises a prophylactically or therapeutically effective amount of the above compound or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof, and a pharmaceutically acceptable carrier.

[0013] The present invention also provides a cosmetic composition, which comprises an effective amount of the above compound or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph thereof, and a cosmetic substrate.

[0014] The present invention also provides a method for extracting the above compound from Forsythia suspensa Vahl, which is characterized by comprising the following steps:

[0015] a) Extraction step: After crushing the dried Forsythia suspensa Vahl, add an alcohol-based aqueous solution with a volume concentration of 30%-95% for extraction, the extraction times are 1-3 times, filter and combine the extraction solutions, and concentrate under reduced pressure until the alcohol smell disappears to obtain a Forsythia suspensa Vahl extract.

[0016] b) Macroporous resin separation step: Load the Forsythia suspensa Vahl extract onto a macroporous resin chromatography column, the resin amount is 1-3 times the mass of the medicinal material, and perform gradient elution successively with water, an ethanol aqueous solution with a volume concentration of 40%-60%, and an ethanol aqueous solution with a volume concentration of 85%-98%. The elution volume of each elution gradient is 4-6 times the column volume; collect the eluate eluted with the 40%-60% ethanol aqueous solution, recover the solvent under reduced pressure to obtain a macroporous resin separation component.

[0017] c) C18 column chromatography separation process: The macroporous resin separated component is loaded onto a C18 chromatography column. The packing material of the C18 chromatography column is octadecylsilyl-bonded silica gel, and the particle size of the packing material is 5 - 100 μm; An acetonitrile-water system is used as the elution solvent, and the elution gradient range is 5% - 70% acetonitrile. The elution method includes gradient elution or isocratic elution; Collect the fractions containing the target component, recover the solvent under reduced pressure to obtain the semi-finished product of cyclopentafulvalenic acid ester;

[0018] d) Gel purification process: The semi-finished product is dissolved and then loaded onto a Sephadex LH-20 gel column. An acetonitrile-water or methanol-water system is used as the elution solvent, and the fractions of cyclopentafulvalenic acid ester with a single purity are collected. After removing the solvent, the pure product is obtained.

[0019] In one embodiment of the present invention, in the above C18 column chromatography separation process, the particle size of the packing material is 5 - 20 μm or 10 - 100 μm, and the elution gradient range is 10% - 40% acetonitrile or 5% - 70% acetonitrile; The macroporous resin is a non-polar macroporous adsorption resin, selected from one or more of D101 resin, AB-8 resin or HP-20 resin; In the extraction process, the alcohol-based aqueous solution is an ethanol aqueous solution, a methanol aqueous solution or a mixed solution thereof, and the extraction methods include room temperature extraction, heating under reflux or ultrasonic-assisted extraction.

[0020] In one embodiment of the present invention, the elution solvent in the above C18 column chromatography separation process is an acetonitrile-water system, and the flow rate of the gradient elution is 0.5 - 5 BV / h; Here, BV means the multiple of the column volume, representing the multiple of the column volume; The elution solvent in the gel purification process is an acetonitrile aqueous solution or a methanol aqueous solution with a volume concentration of 20% - 80%.

[0021] In one embodiment of the present invention, in the above extraction process: after crushing the dried Forsythia suspensa medicinal materials, an alcohol-based aqueous solution with a volume concentration of 30%-95% (preferably 75% ethanol aqueous solution) is added for extraction, and the extraction times are 1-3 times (preferably 2 times). After filtration, the extraction solutions are combined and concentrated under reduced pressure until the alcohol smell disappears to obtain the Forsythia suspensa extract; in the macroporous resin separation process: the Forsythia suspensa extract is loaded onto a macroporous resin chromatography column (the resin amount is 1-3 times the mass of the medicinal materials), and gradient elution is carried out successively with water, an ethanol aqueous solution with a volume concentration of 40%-60% (preferably 50% ethanol aqueous solution), and an ethanol aqueous solution with a volume concentration of 85%-98% (preferably 95% ethanol aqueous solution). The elution volume for each elution gradient is 4-6 times the column volume (preferably 5 times); the eluate of the 40%-60% ethanol aqueous solution is collected, and the solvent is recovered under reduced pressure to obtain the macroporous resin separation component; in the C18 column chromatography separation process: the macroporous resin separation component is loaded onto a C18 chromatography column, and the packing of the C18 chromatography column is octadecylsilyl-bonded silica gel with a packing particle size of 5-100 μm; an acetonitrile-water system is used as the elution solvent, and the elution gradient range is 5%-70% acetonitrile. The elution method includes gradient elution or isocratic elution; the fractions containing the target component are collected, and the solvent is recovered under reduced pressure to obtain the cycloalangionolic acid ester semi-finished product; in the gel purification process: the semi-finished product is dissolved and loaded onto a Sephadex LH-20 gel column, and an acetonitrile-water or methanol-water system is used as the elution solvent. The fractions of cycloalangionolic acid ester with a single purity are collected, and the solvent is removed to obtain the pure product.

[0022] In one embodiment of the present invention, in the above C18 column chromatography separation process, the packing particle size is 5-20 μm or 10-100 μm, and the elution gradient range is 10%-40% acetonitrile or 5%-70% acetonitrile; the macroporous resin is a non-polar macroporous adsorption resin, selected from one or more of D101 resin, AB-8 resin, or HP-20 resin; in the extraction process, the alcohol-based aqueous solution is an ethanol aqueous solution, a methanol aqueous solution, or a mixed solution thereof, and the extraction methods include room temperature leaching, heating under reflux, or ultrasonic-assisted extraction. In one embodiment of the present invention, the elution solvent in the C18 column chromatography separation process is an acetonitrile-water system, and the flow rate of the gradient elution is 0.5-5 BV / h; the elution solvent in the gel purification process is an acetonitrile aqueous solution or a methanol aqueous solution with a volume concentration of 20%-80% (preferably an acetonitrile-water system with a volume concentration of 30%-50%).

[0023] The present invention provides a preparation method of cycloalangionolic acid ester and its application in the preparation of anti-aging drugs, cosmetics, foods, and health products. Description of the Drawings

[0024] Figure 1Structural diagram of the purified cyclofloridoside phosphate ester extracted in Example 1;

[0025] Figure 2 To represent the 1 H- 1 H COSY, HMBC and ROESY key relevant information spectra of the purified cyclofloridoside phosphate ester extracted in Example 1;

[0026] Figure 3 Mass spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (ESI-Positive ion mode);

[0027] Figure 4 Mass spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (ESI-Negative ion mode);

[0028] Figure 5 To represent the 1 1H NMR spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5, 125 MHz);

[0029] Figure 6 To represent the 13 13C NMR spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5, 500 MHz);

[0030] Figure 7 HSQC spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5);

[0031] Figure 8 1H-1H COSY spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5);

[0032] Figure 9 HMBC spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5);

[0033] Figure 10 NOEYS spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1 (Pyridine-d5);

[0034] Figure 11 UV spectrum of the purified cyclofloridoside phosphate ester extracted in Example 1;

[0035] Figure 12 HPLC purity inspection chromatogram of the purified cyclofloridoside phosphate ester extracted in Example 1;

[0036] Figure 13Survival curve of experimental nematodes after extracting and purifying cycloalvioside A in Example 1. Detailed implementation mode

[0037] In this study, a novel compound (cycloalvioside A), a secoiridoid trimer, was isolated and identified from Forsythia suspensa for the first time. Its structure belongs to a compound type not reported in the literature. The compound structure is as follows:

[0038]

[0039] Pharmacological experiments verified the significant anti-aging activity of this compound. In the Caenorhabditis elegans model, cycloalvioside A at 50 μg / mL and 100 μg / mL extended the average lifespan of nematodes by 21.78% and 26.53% respectively (p < 0.001), significantly better than the positive control vitamin E (10 μg / mL, extended by 13.8%, p < 0.01). This result indicates that cycloalvioside A may delay the aging process by regulating the aging-related pathways in nematodes. Combining with the antioxidant mechanism of Forsythia suspensa extract in the knowledge base (such as scavenging free radicals, inhibiting collagen degradation, promoting synthesis, etc.), it is speculated that the activity of this compound may be related to enhancing the antioxidant defense system, inhibiting oxidative stress or regulating metabolic pathways (such as autophagy or mTOR). The molecular structure contains a glycoside unit, and it is speculated that glycosylation modification may be closely related to its anti-aging activity. This discovery not only enriches the research on the chemical constituents of Forsythia suspensa, but also provides a new candidate molecule for anti-aging drug development. The molecular structure of cycloalvioside A contains a glycoside-linked secoiridoid trimer, and glycosylation modification is an important factor affecting the activity of natural products. Existing technical literature studies have shown that the type and number of sugar groups are positively correlated with anti-aging activity (such as the more types of sugar groups in ginsenosides, the stronger the activity); in addition, the position and stability of glycosidic bonds may also affect bioavailability and targeting. Therefore, it is speculated that the glycoside form of this compound may play a role through the following mechanisms: enhanced stability by sugar groups: glycosylation may protect the aglycone from enzymatic or oxidative degradation and prolong its action time in vivo; enhanced targeting: sugar groups may improve the cellular uptake efficiency of the compound by binding to cell surface receptors; synergistic effect: the synergistic effect between the sugar unit and the aglycone may amplify the antioxidant or anti-inflammatory effect.

[0040] In addition, the inventors believe that the secoiridoid trimer of Formula 1 and its glycoside derivatives (such as deglycosylated aglycone or different glycosylation modification forms) may all possess anti-aging activity. In the future, the activity can be further optimized through structural modification (such as changing the sugar group type or the aglycone linkage mode), and its application potential in skin care products or drugs can be explored. In the present invention, Glc represents glucosyl, Glc represents glucosyl, Fru represents fructosyl, Gal represents galactosyl, Man represents mannosyl, and Xyl represents xylosyl.

[0041] The cyclic pentaerythrin esters in the present invention can be prepared by the following typical technical methods:

[0042] Extraction process: After the dried Forsythia suspensa medicinal material is crushed, an alcohol aqueous solution is added, extracted twice, filtered, the two extracts are combined, and the decompression is reduced until there is no alcohol taste to obtain the Forsythia suspensa extract;

[0043] Macroporous resin separation process: Load the Forsythia suspensa extract onto a macroporous resin chromatography column (the amount of resin is 1-2 times the amount of the medicinal material), and elute with water, 50% ethanol water, and 95% ethanol water in sequence, with each gradient elution being 5 times the column volume. Collect the 50% ethanol water eluate and recover the solvent under reduced pressure;

[0044] Medium-pressure C18 column separation step: The 50% ethanol water elution fraction of the macroporous resin is loaded onto a medium-pressure C18 chromatography column, and gradient eluted with an acetonitrile water system (5%→70%), and the target fractions are collected, combined, and the solvent is recovered under reduced pressure to obtain a fraction containing pentasperglycans;

[0045] High-pressure C18 column separation step: subjecting the pentaerythritol ester-containing components to high-pressure preparation, eluting with acetonitrile formic acid water, collecting the target peak, and recovering the solvent under reduced pressure to obtain the semi-finished product of cyclopentaerythritol ester;

[0046] Gel purification process: After dissolving the semi-finished product of cyclopentaerythritol ester, apply it to Sephadex LH-20 gel column, elute with acetonitrile water, collect the single cyclopentaerythritol ester fraction, remove the solvent to obtain the pure cyclopentaerythritol ester.

[0047] In one embodiment of the present invention, in the extraction step, 0%-100% ethanol aqueous solution or methanol aqueous solution is used as the alcohol aqueous solution for extraction, and preferably 75% ethanol aqueous solution is used for extraction in the extraction step.

[0048] In one embodiment of the present invention, in the extraction step, the extraction method includes room temperature extraction, heating extraction or reflux extraction.

[0049] In one embodiment of the present invention, in the macroporous resin separation step, the selected macroporous resin is a non-polar macroporous adsorption resin, including D101 resin, AB-8 resin, HP20 resin and the like.

[0050] In one embodiment of the present invention, in the medium pressure C18 column separation process, the selected filler type is octadecylsilane bonded silica gel, and the filler particle size is in the range of 10-100 μm; the elution solvent used is 5%-70% acetonitrile aqueous solution, and the elution mode includes elution elution and isocratic elution.

[0051] In one embodiment of the present invention, in the high-pressure C18 column separation process, the selected packing type is octadecylsilyl-bonded silica gel, and the particle size of the packing is in the range of 5-20 μm; the elution solvent used is an aqueous solution of 10%-40% acetonitrile, and the elution methods include gradient elution and isocratic elution.

[0052] In one embodiment of the present invention, in the gel purification process, the gel used is Sephadex LH20; the elution solvents used are methanol-water and acetonitrile-water solvent systems.

[0053] The cyclofloridoside acid ester obtained in the present invention has an anti-aging effect. It has the effect of prolonging the lifespan of Caenorhabditis elegans and can be used to prepare anti-aging food or pharmaceutical compositions, which include cyclofloridoside acid ester as an active ingredient and excipients. The excipients are well-known to those skilled in the art, such as excipients, fillers, binders, disintegrants, wetting agents, surfactants, etc.

[0054] The following further elaborates on this application in conjunction with examples.

[0055] Example 1 Extraction and Purification

[0056] Herbal medicine extraction: 10 kg of Forsythia suspensa herbs are crushed and extracted with 100 L of 75% ethanol at room temperature for 3 days. After filtration, the medicinal residues are extracted with another 100 L of 75% ethanol at room temperature for 3 days. After filtration, the two extraction solutions are combined and concentrated under reduced pressure at 55°C until the alcohol smell disappears (about 20 L) to obtain the total Forsythia suspensa extract.

[0057] Macroporous resin separation: 10 kg of D101 macroporous resin is soaked in ethanol overnight and then washed with water until the alcohol is removed, and then loaded into a chromatography column (230*1180 mm). The total Forsythia suspensa extract is loaded onto the D101 resin chromatography column. After loading, it is eluted successively with water, 50% ethanol-water, and 95% ethanol-water, with 150 L eluted for each gradient. The 50% ethanol-water eluate is collected and concentrated under reduced pressure at 55°C to obtain the 50% ethanol-water elution fraction of the resin.

[0058] Medium-pressure C18 separation: The 50% ethanol-water elution fraction of the resin is filtered and loaded onto a medium-pressure C18 chromatography column (260*600 mm), and eluted successively with 5%, 15%, 25%, 50%, and 70% acetonitrile-water gradients, with 100 L eluted for each gradient. One bottle is collected every 5 L, and the target is detected by HPLC. The fractions containing the target component are combined and concentrated under reduced pressure at 55°C to obtain the fraction containing cyclofloridoside acid ester.

[0059] High-pressure preparation: Inject the component containing cycloalvioside into high-pressure liquid chromatography for preparation (Daiso ODS-RPS 5μm, 50*250mm), elute with 30% acetonitrile formic acid water, detect at a wavelength of 230nm, flow rate of 60mL / min, collect the target peak, and obtain the semi-finished product of cycloalvioside after concentration under reduced pressure.

[0060] Gel purification: Dissolve the semi-finished product of cycloalvioside obtained by high-pressure preparation and perform gel column separation and purification. After the sample is loaded onto the gel column (25*1200mm, Sephadex LH-20), it is eluted isocratically with 10% acetonitrile water, and one bottle is collected every 45mL. After detecting the purity of each fraction by HPLC, the fractions with qualified purity are combined, concentrated under reduced pressure, and freeze-dried to obtain 1484mg of cycloalvioside product, with a yield of 0.0148%. The HPLC detection results of the product are shown in the appendix Figure 12 。

[0061] Example 2 Structure Identification

[0062] The cycloalvioside sample is a white powder, and HR-ESI-MS shows that m / z is 1073.3751 [M-H] - (calculated value 1073.3713), and the molecular formula is C 48 H 66 O 27 。

[0063] 13 The 13C NMR spectrum (see Table 1 in the appendix) shows that the compound has a total of 16 carbon signals, and it is speculated that its structure is composed of 3 identical structural units, each containing 16 carbons. 1 The 1H NMR spectrum (see Table 1 in the appendix) shows 1 vinylic proton signal δ H 7.82(1H, s, H-3), 1 acetal proton signal δ H 5.36(1H, d, J = 7.3Hz, H-1), 1 anomeric proton signal of sugar δ H 5.34(1H, d, J = 8.0Hz, H-G1); 13 The 13C NMR spectrum shows 2 vinylic carbon signals δ C 153.3 and 111.9, 1 ester carbonyl signal δ C 167.9 and 1 anomeric carbon signal δ C 101.3; The above are all characteristic signals of iridoid glycosides, so it is speculated that the 16-carbon structural unit is an iridoid glycoside.

[0064] Based on the carbon spectrum data and the coupling constant (8.0Hz) of the anomeric proton of the sugar, it is inferred that the sugar moiety in this compound is β-D-glucopyranose. According to 1 H- 1In the H-COSY spectrum, H-5 is correlated with H2-6 and H-9, H-8 is correlated with H2-7 and H-9, and H2-7 is correlated with H2-6, from which a five-membered ring can be deduced; in the HMBC spectrum, the correlation between H-1 and C-3, C-G1 indicates that the sugar unit is attached to C-1 of the iridoid aglycone. In the ROESY spectrum, the correlation between H-1 and H-8, and between H-5 and H-9 shows that both H-1 and H-8 are of the α-type, and both H-5 and H-9 are of the β-type. The above NMR data and the stereoconfiguration of the chiral centers are consistent with the literature [1] reported adoxosidic acid, indicating that this 16-carbon structural unit is adoxosidic acid. The molecular formula of the compound is C 48 H 66 O 27 , with an unsaturation degree of 16. Among them, 15 degrees of unsaturation come from 3 adoxosidic acid structural units. Combining with 13 the 13C NMR data, it is speculated that the remaining 1 degree of unsaturation comes from the ring formation of the 3 structural units. In the HMBC spectrum, the correlation between H2-10 and C-11 indicates that C-11 and C-10 are linked by an ester bond to form a ring. 1 1H and 13 13C NMR spectra show only one set of signals for adoxosidic acid. In summary, the compound has a cyclic trimer structure as shown in Appendix Figure 1 , which is reported for the first time in this application and is named Cycloadoxosidate.

[0065] Table 1 is a relational diagram of the NMR data attribution of the cycloadoxosidic acid extracted and purified in Example 1;

[0066] Table 1 NMR Data Attribution of Cycloadoxosidic Acid

[0067]

[0068] Data were recorded in Pyrindine-d5, at 500MHz and 125MHz, respectively.

[0069] Example 3 Pharmacological Activity Test

[0070] The anti-aging activity test was carried out using the cycloadoxosidate product obtained in Example 1:

[0071] Synchronization of nematodes: Before the nematodes grow into adults and start laying eggs but have not laid a large number of eggs, a row of bright-colored eggs can be observed in the abdominal cavity of the nematodes under a microscope. Rinse the surface of the NGM agar plate with M9 buffer, collect the nematodes into an EP tube, let it stand for 1-2 minutes until the nematodes sink to the bottom, discard the supernatant. Rinse three times with M9 buffer according to this method, and finally collect the nematodes into a new EP tube. Add Bleach solution (200 μL), shake and lyse for 7 minutes, centrifuge at 5600 r for 1 minute, discard the supernatant, add M9 to wash and centrifuge. After centrifugation, add M9 buffer, incubate at 20 °C for about 10-12 hours and then transfer to an NGM culture plate. L4-stage nematodes can be obtained after 60-72 hours.

[0072] Preparation of M9 buffer: 0.3 g KH2PO4, 1.512 g Na2HPO4·12H2O, 0.6 g NaCl, 0.1 mL MgSO4 (1 M), make up the volume to 100 mL with distilled water.

[0073] Preparation of Bleach solution: 20 mL 100% NaClO, 25 mL NaOH (2 mol / L), 55 mL double-distilled water.

[0074] Drug addition: The composition of the solution in each well of the nematode 96-well plate liquid medium: 121.2 μL M9 + 13.47 μL OP50 + 0.3 μL FUdR (12.5 M) + 15 μL cyclovirobuxine sample (1 mg / mL), with a total volume of 150 μL. After adding the above solution to each well, pick in the nematodes.

[0075] Observation of nematodes: Observe the life and death of nematodes and count (gently touch the nematodes with a nematode needle, and those that do not wriggle are dead), and continue to observe until the last nematode dies. Draw a survival curve and compare the differences. The data uses python3.10 and the corresponding libraries lifelines == 0.27.8, matpplotlib == 3.6.0, pandas == 1.4.2, numpy == 1.22.4, and the Kaplan-Meier method is used for survival curve analysis. p < 0.05 is marked as "*", which is of significant significance; p < 0.01 is marked as "**", which is of extremely significant significance.

[0076] The experimental results show that compared with the blank solvent control group, 50 μg / mL of cyclovirobuxine can increase the average survival life of nematodes from 19.6 ± 4.68 days to 23.8 ± 5.87 days, extending the life cycle by 21.78% (p < 0.001); 100 μg / mL of cyclovirobuxine can increase the average life of nematodes from 19.6 ± 4.68 days to 24.8 ± 5.75 days, extending the life cycle by 26.53% (p < 0.001). The survival curve is shown in the appendix Figure 13 。

[0077] In summary, the above embodiments are only preferred embodiments of the present invention and are not intended to limit the protection scope of the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A iridoid compound represented by Formula 1 or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof, wherein each X independently represents O, S or CH2, and each R independently represents H, Glc, Fru, Gal or Xyl.

2. The compound according to claim 1, or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof, wherein, X represents O, and R represents H or Glc.

3. The compound according to claim 1, or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof, wherein, The compound represented by Formula 1 has the following structure, 。 4. Use of the compound according to any one of claims 1 to 3 in the preparation of a drug for anti-aging.

5. A pharmaceutical composition comprising a prophylactically or therapeutically effective amount of the compound according to any one of claims 1 - 3 or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and isotope-labeled compound thereof, and a pharmaceutically acceptable carrier.

6. A cosmetic composition comprising a prophylactically or therapeutically effective amount of the compound according to any one of claims 1 - 3 or a pharmaceutically acceptable salt, optical isomer, stereoisomer, polymorph, and a cosmetic base.

7. A method for extracting the compound according to claim 3 from Forsythia suspensa, characterized in that, Comprising the following steps: a) Extraction step: After pulverizing the dried Forsythia suspensa medicinal material, add an alcohol-based aqueous solution with a volume concentration of 30% - 95% for extraction, the number of extraction times is 1 - 3 times, filter and combine the extraction solutions, and concentrate under reduced pressure until the alcohol smell disappears to obtain a Forsythia suspensa extract; b) Macroporous resin separation step: Load the Forsythia suspensa extract onto a macroporous resin chromatography column, the resin amount is 1 - 3 times the mass of the medicinal material, and perform gradient elution successively with water, an ethanol aqueous solution with a volume concentration of 40% - 60%, and an ethanol aqueous solution with a volume concentration of 85% - 98%. The elution volume for each elution gradient is 4 - 6 times the column volume; Collect the eluate of the 40% - 60% ethanol aqueous solution, recover the solvent under reduced pressure to obtain a macroporous resin separation component; c) C18 column chromatography separation step: Load the macroporous resin separation component onto a C18 chromatography column, the packing of the C18 chromatography column is octadecylsilyl-bonded silica gel, and the packing particle size is 5 - 100 μm; Use an acetonitrile - water system as the elution solvent, and the elution gradient range is 5% - 70% acetonitrile. The elution method includes gradient elution or isocratic elution; Collect the fraction containing the target component, recover the solvent under reduced pressure to obtain a semi-finished product of cycloalvulinic acid ester; d) Gel purification step: Dissolve the semi-finished product and load it onto a Sephadex LH-20 gel column, use an acetonitrile - water or methanol - water system as the elution solvent, collect the fraction of cycloalvulinic acid ester with a single purity, and remove the solvent to obtain a pure product.

8. The method according to claim 7, characterized in that: i) In the C18 column chromatography separation step, the packing particle size is 5 - 20 μm or 10 - 100 μm, and the elution gradient range is 10% - 40% acetonitrile or 5% - 70% acetonitrile; ii) The macroporous resin is a non-polar macroporous adsorption resin, selected from one or more of D101 resin, AB-8 resin or HP-20 resin; iii) In the extraction process, the alcohol-based aqueous solution is an ethanol aqueous solution, a methanol aqueous solution or a mixed solution thereof, and the extraction methods include extraction at room temperature, heating under reflux or ultrasonic-assisted extraction.

9. The method according to claim 7 or 8, characterized in that: a) The elution solvent for the C18 column chromatography separation process is an acetonitrile-water system, and the flow rate of the gradient elution is 0.5-5 BV / h; b) The elution solvent for the gel purification process is an acetonitrile aqueous solution or a methanol aqueous solution with a volume concentration of 20%-80%.

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