Use of polygonatum extract in preparation of anti-aging drugs

Abstract

The application discloses application of a polygonatum officinale extract in preparation of anti-aging drugs and belongs to the technical field of traditional Chinese medicines. The application fully studies the anti-aging mechanism of the polygonatum officinale extract, and it is clear that the polygonatum officinale extract can be used for improving organ injury, aging-related secretory phenotype and intestinal flora imbalance and the like, thereby providing a scientific basis for anti-aging application of the polygonatum officinale extract and providing a reference for further development of the polygonatum officinale extract.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine technology, and more specifically to the application of Polygonatum odoratum extract in the preparation of anti-aging drugs. Background Technology

[0002] Polygonatum odoratum (Mill.) Druce, a perennial herb belonging to the genus Polygonatum in the family Liliaceae, was first recorded in the *Shennong Bencao Jing* (Shennong's Classic of Materia Medica), where it was known as "Nüwei" and "Weirui," and was listed as a superior-grade herb. Polygonatum odoratum was one of the first Chinese medicinal herbs included in the list of herbs that are both food and medicine. With the development of traditional Chinese medicine and the health industry in my country, its pharmacological and health-promoting value has received increasing attention, and it has been extensively studied by numerous medical and food scientists.

[0003] Aging refers to the physiological process in which the body's physiological and psychological adaptability to the environment progressively declines, gradually leading to death. With the aging of modern society, aging has become a significant social issue. Traditional Chinese medicine, with its holistic health concept of "prevention before aging," has become a popular choice for many seeking to prolong life. Furthermore, many medicinal and edible herbs serve as sources of anti-aging drugs.

[0004] However, current research on the anti-aging effects and mechanisms of Polygonatum odoratum is insufficient, and relevant information is relatively scarce. Therefore, it is necessary to utilize modern analytical methods to explore the mechanisms of action of Polygonatum odoratum in anti-aging applications, uncovering mechanistic pathways from multiple perspectives and dimensions. This will provide a scientific basis for the anti-aging applications of Polygonatum odoratum and offer a reference for its further development. Thus, investigating the uses and mechanisms of action of Polygonatum odoratum in anti-aging drugs is a problem that urgently needs to be solved by those skilled in the art. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide an application of Polygonatum odoratum extract in the preparation of anti-aging drugs.

[0006] Furthermore, Polygonatum odoratum extract is used to improve organ damage, age-related secretory phenotypes, and gut microbiota dysbiosis; Polygonatum odoratum extract significantly reduces the levels of IL-1β, MCP-1, and GDF-15 in serum SASP; Polygonatum odoratum extract reduces the dysbiosis index and regulates the gut microbiota structure.

[0007] Furthermore, Polygonatum odoratum extract was used to upregulate Sirt1 protein expression and downregulate P16 protein expression in the spleen.

[0008] Furthermore, Polygonatum odoratum extract can be used to improve the composition of spleen immune cell subsets and age-related metabolic phenotypes.

[0009] Furthermore, Polygonatum odoratum extract significantly increased the number of naive CD4+ T cells, decreased the number of memory CD8+ T cells, and reduced the expression of P16 mRNA in spleen T cells.

[0010] Furthermore, Polygonatum odoratum extract improves CD4+ T cell metabolism and delays aging by regulating the Park7 / Sirt1 signaling pathway.

[0011] Furthermore, Polygonatum odoratum extract was used to delay yeast senescence and upregulate yeast Sir2 gene expression.

[0012] The Solomon's seal extract of this invention is preferably a water extract of Solomon's seal.

[0013] The preparation method of the Polygonatum odoratum extract of the present invention includes the following steps: taking Polygonatum odoratum and decocting it with water 1-4 times, each time for 1-4 hours, centrifuging and filtering the decoction, and concentrating the filtrate. Further, the preferred method for preparing the water extract of Polygonatum odoratum includes the following steps: taking Polygonatum odoratum and decocting it with water three times (5-6 times the amount, 4-5 times the amount, and 4-5 times the amount), the first time for 3 hours, the second and third times for 2 hours each, centrifuging and filtering the decoction, and concentrating the filtrate to a clear extract with a relative density of 1.10-1.20 (60℃~70℃).

[0014] The Polygonatum odoratum extract of the present invention, with the addition of excipients, is formulated into a clinically acceptable anti-aging pharmaceutical preparation.

[0015] The beneficial effects of this invention are as follows: This invention elucidates the process by which Polygonatum odoratum extract regulates immune response and delays aging through multiple pathways, and clarifies that CD4+ T cells are the key immune cells for Polygonatum odoratum extract to regulate immunity and delay aging. Polygonatum odoratum extract can be used to prepare anti-aging drugs. Polygonatum odoratum extract can be mixed with excipients to prepare oral liquids, ointments, tablets, capsules, pills or granules, etc. These preparations are convenient to take and can be taken alone or in combination with other drugs. Attached Figure Description

[0016] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0017] Figure 1 To investigate the effect of Polygonatum odoratum extract on the phenotypes of aged mice on a normal diet. (A) Experimental flowchart; (BD) Organ index; (FJ) SASP; (LM) Tight junction protein of the intestinal barrier; (O) Gut microbiota alpha diversity index; (P) PCoA analysis of gut microbiota beta diversity.

[0018] Figure 2 To investigate how Polygonatum odoratum extract improves phenotypes in aging mice on a normal diet. (A) Intestinal flora dysbiosis index; (B) Firmicutes / Bacteroidetes ratio; (C) Genus-level relative abundance; (DI) p16 mRNA expression as an aging marker in various organs; (JK) p16 protein expression in the spleen; (LN) Sirt1 protein expression in the spleen.

[0019] Figure 3 To investigate how Polygonatum odoratum extract improves phenotypes associated with aging mice on a high-fat diet. (A) Experimental flowchart; (BH) Organ index; (IK) SASP; (LM) Lung function.

[0020] Figure 4 To investigate how Polygonatum odoratum extract improves phenotypes associated with a high-fat diet in aging mice. (AB) Gut microbiota alpha diversity index; (C) Gut microbiota beta diversity PCoA analysis; (D) Gut microbiota dysbiosis index; (E) Firmicutes / Bacteroidetes ratio; (F) Phylum-level relative abundance; (G) Genus-level relative abundance; (HK) Abundance of bacteria with the same regulatory trend after Polygonatum odoratum extract intervention under normal and high-fat diet conditions.

[0021] Figure 5 To investigate the effect of Polygonatum odoratum extract on the number of immune cells in aging mice on a normal diet. (AF) Peripheral blood immune cell count; (GH) Splenic B cell subset composition; (KQ) Splenic T cell subset composition; (RU) Splenic T cell aging marker mRNA expression.

[0022] Figure 6 To investigate how Polygonatum odoratum extract improves the number of immune cells in aging mice on a high-fat diet. (AF) Peripheral blood immune cell count; (GH) Splenic B cell subset composition; (JQ) Splenic T cell subset composition.

[0023] Figure 7 Polygonatum odoratum extract improves CD4 levels in aging mice with normal diet + T cell senescence and metabolic phenotype. (AD) Protein expression of Sirt1, HUR, P16, and β-actin; (E) Lactate content in mouse blood samples; (F) Lactate content in mouse spleen samples; (G) CD4 + T cell lactate content; (HN)CD4 + T-cell energy metabolism phenotype.

[0024] Figure 8 Proteomics analysis of the regulation of CD4 in aging mice by Polygonatum odoratum extract on normal diet +Differential protein screening and functional classification in T cells. (A) Volcano plot of differentially expressed proteins in Control vs. Model; (B) Volcano plot of differentially expressed proteins in Model vs. Pol; (C) Secondary classification of differentially expressed proteins in GO; (D) Cluster analysis of differentially expressed proteins.

[0025] Figure 9 To suppress Park7's effect on CD4 + Effects of T cell senescence and energy metabolism phenotype. (AD) mouse CD4 + Proteins of Slc38a1, PD-1, Park7, and β-actin in T cells; (EF) inhibits Park7's effect on CD4. + The effect of Sirt1 protein in T cells; (G) inhibition of Park7 on CD4 + Effects of T cell lactate content; (HN) inhibition of Park7 on CD4 + The influence of T cell energy metabolism phenotype.

[0026] Figure 10 The growth curves of Solomon's seal extract on yeast are shown in (a) and (b) for the effects of 10 μg / mL-100 mg / mL Solomon's seal extract on yeast.

[0027] Figure 11 The effect of Polygonatum odoratum extract on yeast survival rate: (a) Effect of Polygonatum odoratum extract on yeast survival rate; (b) Viable cell plate count.

[0028] Figure 12 The effects of Polygonatum odoratum extract, rutin, and methyl ophiopogon flavanone B on yeast survival rate.

[0029] Figure 13 Images of Lü's basic methylene blue staining (blank group and different doses of Polygonatum odoratum extract).

[0030] Figure 14 For Lüssler basic methylene blue staining (blank group and rutin (0.1 mg / mL) and methyl ophiopogon flavanone B (0.1 mg / mL) group).

[0031] Figure 15 The effect of Polygonatum odoratum extract on Sir2 DNA expression in yeast cells (Control group, Model group, and different doses of Polygonatum odoratum extract). ## P<0.01vs.Control; *P<0.05, **P<0.01, ***P<0.001vs.Model, n=3).

[0032] Figure 16The effect of Polygonatum odoratum extract on Sir2 DNA expression in yeast cells (Control group, different doses of rutin, and different doses of methyl ophiopogon flavanone B) (**P<0.01 vs. Control, n=3). Detailed Implementation

[0033] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] Example 1

[0035] This invention provides a method for preparing Polygonatum odoratum extract:

[0036] Take 10kg of Polygonatum odoratum slices, add water and decoct three times (5 times, 4 times, and 4 times the amount), the first time for 3 hours, the second and third times for 2 hours each, centrifuge and filter the decoction, and concentrate the filtrate under reduced pressure to a clear extract with a relative density of 1.10-1.20 (60℃~70℃).

[0037] Example 2

[0038] This invention provides a method for preparing Polygonatum odoratum extract:

[0039] Take 10 kg of Polygonatum odoratum slices, add water and decoct twice (6 times the amount and 5 times the amount), the first time for 4 hours and the second time for 2 hours. Centrifuge and filter the decoction, and concentrate the filtrate under reduced pressure to a clear extract with a relative density of 1.10-1.20 (60℃~70℃).

[0040] Example 3

[0041] This invention provides a method for preparing Polygonatum odoratum extract:

[0042] Take 10kg of Polygonatum odoratum slices, add water and decoct three times (6 times, 5 times, and 5 times the amount), the first time for 3 hours, the second and third times for 2 hours each. Centrifuge and filter the decoction, and concentrate the filtrate under reduced pressure to a clear extract with a relative density of 1.10-1.20 (60℃~70℃).

[0043] The experimental grouping method, preparation of the drug group, and administration method are as follows in the following examples:

[0044] Grouping method: 3-month-old C57BL / 6J mice were used as the control group. 17-month-old mice were randomly divided into 5 groups: Model group, medium-dose Polygonatum odoratum group (Pol-M), high-dose Polygonatum odoratum group (Pol-H), and positive control drug rapamycin group (Rap), with 8 mice in each group.

[0045] Preparation of different dosage groups of Polygonatum odoratum extract:

[0046] The preparation process of Polygonatum odoratum extract is as follows: Polygonatum odoratum was extracted three times with water. The first extraction was carried out with 5 times the amount of water for 3 hours, and the second and third extractions were carried out with 4 times the amount of water for 2 hours each. The extract was centrifuged and filtered for later use. It was then concentrated in an open container to a relative density of 1.17 (60℃). After being refrigerated and allowed to stand for 16 hours, it was filtered through a 200-mesh filter, and 3.143 kg of extract was collected with a relative density of 1.19 (20℃).

[0047] Medium dose of Polygonatum odoratum (Pol-M): Weigh 0.7g and dissolve in 10mL of pure water to prepare a 70mg / mL medium dose Pol solution.

[0048] High dose of Polygonatum odoratum (Pol-H): Weigh 1.4g and dissolve in 10mL of pure water to prepare a 140mg / mL high dose Pol solution.

[0049] Rapamycin (Rap): Weigh 4 mg and add 1 mL of anhydrous ethanol, 0.5 mL of PEG-400 and 0.5 mL of L-Ween-80 to dissolve it. Make up the volume to 10 mL with pure water and use an ultrasonic cleaner for 30 min to aid dissolution. Prepare a 0.4 mg / mL rapamycin solution.

[0050] Administration methods: Except for the rapamycin group, which was administered intraperitoneally every other day, all other groups of mice were administered the drug via gavage once daily for 8 consecutive weeks. Based on the clinical dosage and the conversion between mouse and human dosages (pharmacopoeia dosage: 6-12 g / day), medium- and high-dose groups of Polygonatum odoratum were established. The medium-dose group (0.7 g / kg) was approximately equal to the clinical equivalent dose (1.56 g / kg of raw drug in mice), and the high-dose group (1.4 g / kg administered via gavage) was approximately twice the clinical equivalent dose (3.12 g / kg of raw drug in mice). The rapamycin group was administered intraperitoneally at 4 mg / kg, while the control and model groups received the same volume of solvent. Mice were weighed every two days, and the dosage volume was adjusted promptly based on weight changes.

[0051] Example 4: Study on the effect of Polygonatum odoratum extract on improving aging and related phenotypes

[0052] Methods: Seventeen-month-old male C57BL / 6J mice were used as an aging model. They were administered Polygonatum odoratum extract (water extract) / rapamycin (positive control) for 8 weeks under normal and high-fat diet conditions, respectively. Histopathological staining, double-antibody sandwich enzyme-linked immunosorbent assay (ELISA), qPCR, Western blotting, and gut microbiota diversity analysis were used to detect the effects of Polygonatum odoratum extract (Pol) on aging-related phenotypes in mice, including organ pathological damage, organ aging markers, serum senescence-associated secretory phenotypes (SASPs) such as monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-α (TNF-α), osteopontin (OPN), interleukin-1β (IL-1β), activin A (Activin A), growth differentiation factor-15 (GDF-15), and gut microbiota dysbiosis, to evaluate the anti-aging effect of Polygonatum odoratum extract.

[0053] Results: Combined with appendix Figure 1-4 Polygonatum odoratum extract significantly improved organ damage, saline splenic acid saturates (SASP), and gut microbiota dysbiosis in aging mice. Under normal dietary conditions, Polygonatum odoratum extract tended to downregulate the expression of the aging marker P16 in various organs and tissues, and improved the organ indices of the heart, kidneys, and spleen, as well as pathological damage to the lungs and intestines in aging mice. Specifically, Polygonatum odoratum extract improved alveolar dilatation, widened alveolar spaces, increased the expression of intestinal atresia zona 1 (ZO-1) and atresia protein (Occludin), and significantly reduced the levels of IL-1β, MCP-1, and GDF-15 in serum SASP of aging mice; simultaneously, it reduced the gut microbiota dysbiosis index and regulated the gut microbiota structure. Under high-fat dietary conditions, Polygonatum odoratum extract reduced the spleen index, improved lung function in aging mice, had no significant effect on serum SASP, and similarly reduced the gut microbiota dysbiosis index and regulated the gut microbiota structure. Furthermore, the abundance of bacteria exhibiting similar regulatory trends under both normal and high-fat diets were screened using Polygonatum odoratum extract. Upregulated bacteria included Bifidobacteriaceae, Enterobacter, and Lactobacillus johnsonii, while downregulated bacteria included unclassified Helicobacter. By examining the effects of Polygonatum odoratum extract on aging markers in different tissues of aging mice, it was found that Polygonatum odoratum extract significantly reduced the expression of P16 protein in the spleen of aging mice and upregulated the expression of Sirt1 protein in the spleen. These results suggest that Polygonatum odoratum extract may exert its anti-aging effect by influencing the senescence of immune cells in the spleen.

[0054] The results showed that Polygonatum odoratum extract has the effect of delaying the aging phenotype. A high-fat diet further exacerbates the aging phenotype and has some damage to the immune system. The intervention effect of Polygonatum odoratum extract on naturally aging mice under a high-fat diet was examined and compared with the effect of calorie restriction intervention. The results showed that Polygonatum odoratum extract significantly reduced the spleen index of aging mice, consistent with the results of the calorie restriction group; Polygonatum odoratum extract also improved lung function and gut microbiota imbalance, but the intervention effect was not as significant as that of a normal diet, suggesting that dietary conditions should be considered when administering the drug clinically.

[0055] Example 5: Study on the effect of Polygonatum odoratum extract on improving the composition of spleen immune cell subsets and aging-related metabolic phenotypes.

[0056] Methods: Flow cytometry was used to detect the effects of Polygonatum odoratum extract (water extract) on the composition of immune cell subsets in peripheral blood and spleen of aging mice, thereby exploring the regulatory role of Polygonatum odoratum extract in immune function during aging and identifying the main immune cell types involved. Subsequently, primary aging immune cells were sorted using magnetic bead sorting and cultured in vitro. Polygonatum odoratum-containing serum was prepared and used for intervention. Biochemical assays, a Seahorse cell energy metabolism analyzer, and Western blotting were used to detect the effects of Polygonatum odoratum extract on the metabolic phenotype of immune cells, including lactate content, cellular energy metabolism, and Sirtuin1 (Sirt1) protein expression, to evaluate the regulatory effect of Polygonatum odoratum extract on immune aging.

[0057] Result: Combination Figure 5-7 Based on the established understanding that the spleen is a key organ regulated by Polygonatum odoratum extract in the process of delaying aging, this study investigated the regulatory effect of Polygonatum odoratum extract on the immune function of aging mice. The results showed that Polygonatum odoratum extract had no significant effect on the composition of B cell subsets in the peripheral blood and spleen of aging mice on a normal diet, but it significantly increased the amount of naive CD4+ cells in the spleen. + T cell count, reduced memory (TEM) CD8 + It reduced the number of T cells and decreased the expression of p16 mRNA in splenic T cells. However, under a high-fat diet, Polygonatum odoratum extract significantly increased naive CD4+ in the spleen of aging mice. + T cell count. CD4+ count in sorted spleen. + Western blot analysis of T cells revealed that Polygonatum odoratum extract significantly upregulated Sirt1 protein expression and downregulated P16 protein expression, suggesting that the main anti-aging effect of Polygonatum odoratum extract is on spleen CD4 immune cells. + T cells. Metabolic phenotype analysis revealed that Polygonatum odoratum extract could reduce lactate levels in the peripheral blood and spleen of aging mice and improve primary CD4+ in aging mice. +The levels of T cell non-mitochondrial oxygen consumption, basal oxidative phosphorylation, proton leakage, and glycolysis were measured, indicating that Polygonatum odoratum extract can upregulate CD4. + Sirt1 expression in T cells improves cell metabolism and plays a role in delaying aging.

[0058] Given the crucial role of Sirt1 in the metabolism of senescent cells, regulating Sirt1 expression can not only affect the metabolic function of immune cells, including T cells, but alterations in metabolic phenotype can also conversely regulate Sirt1 expression; for example, upregulation of lactate levels inhibits Sirt1 expression. Further, the effects of Polygonatum odoratum extract on CD4... + The study examined changes in T cell metabolic phenotypes and found that Polygonatum odoratum extract could reduce lactate levels in peripheral blood and spleen tissue of aging mice, and also reduce aging CD4 counts at the in vitro cellular level. + T cell lactate levels and improved energy metabolism, including improved CD4 levels. + The study also investigated the levels of T cell non-mitochondrial oxygen consumption, basal oxidative phosphorylation, proton leakage, and glycolysis. These results indicate that Polygonatum odoratum extract can promote Sirt1 expression and improve CD4 levels. + T cell metabolism thus delays CD4 metabolism. + T-cell senescence.

[0059] Example 6: Polygonatum odoratum extract improves spleen CD4 levels by regulating the Park7 / Sirt1 signaling pathway. + Research on the mechanism of T cell metabolism in delaying aging ( Figure 8-9 )

[0060] Methods: Microproteomics was used to detect, screen, and predict differentially expressed proteins that play a key role in the expression of Sirt1 protein on target immune cells by Polygonatum odoratum extract (water extract), and the differentially expressed proteins in vivo were verified by Western blotting. In vitro aging CD4... + In a T-cell model, protein inhibitor intervention was used. Biochemical assays, a Seahorse cell energy metabolism analyzer, and Western blotting were employed to detect the inhibitor's effect on improving CD4 counts using Polygonatum odoratum extract. + This study investigated the effects of Polygonatum odoratum extract on T cell metabolic phenotypes, including lactate content, cellular energy metabolism, and Sirt1 protein expression, to explore the mechanism by which it improves immune cell metabolism and delays aging by regulating the Sirt1 signaling pathway.

[0061] Result: Combination Figure 8-9 Spleen CD4+ T-cell proteomics results revealed 62 differentially expressed proteins with a retrograde effect after administration of Polygonatum odoratum extract. Literature review identified GATA3, Slc38a1, PD-1, PARK7, and Oma1 as metabolic-related differentially expressed proteins, and Eomes, Thrombomomodulin, PD-1, and PARK7 as differentially expressed proteins associated with Sirt1. Therefore, PD-1 and PARK7 were selected for validation. Slc38a1, associated with glycolysis, was also selected for validation. Western blotting validated the selected differentially expressed proteins. Compared to the Model group, both the Pol-M and Pol-H groups significantly reduced PD-1 protein expression (P<0.05, P<0.001), while the Pol-H group increased Park7 protein expression (P<0.01). It is speculated that Park7 may be a key factor in the delayed CD4+ retardation effect of Polygonatum odoratum extract. + The target of T cell senescence was investigated. Park7 expression was inhibited using tetramethylthiuram disulfide, and the effects of Polygonatum odoratum extract on senescent primary CD4+ were then observed. + The effects of T cell metabolic phenotype: Compared with the control group, Sirt1 protein expression in senescent primary CD4+ T cells was significantly decreased (P<0.001), while Sirt1 protein expression in primary CD4+ T cells in the drug-containing serum intervention group was significantly increased (P<0.001). However, after the addition of Park7 inhibitor, Sirt1 protein expression in CD4+ T cells was significantly decreased (P<0.001), and there was no significant difference compared with the model group. This indicates that the addition of Park7 inhibitor interfered with the activation of CD4+ T cells by Polygonatum odoratum extract. + Inhibition of Sirt1 protein expression in T cells leads to downregulation of Sirt1 protein expression. Simultaneously, inhibition of Park7 also affects CD4 expression. + T cell non-mitochondrial oxygen consumption, basal oxidative phosphorylation level, ATP production, and glycolysis level were significantly decreased, indicating that Park7 is a regulator of CD4 by Polygonatum odoratum extract. + Key proteins involved in T cell Sirt1 expression and metabolism, further clarifying that Polygonatum odoratum extract improves CD4 expression by regulating the Park7 / Sirt1 signaling pathway. + The role of T-cell metabolism in delaying aging.

[0062] Example 7: Polygonatum odoratum extract, active compound rutin, and methyl ophiopogon flavanone B improve spleen CD4 + Research on the role of T cell metabolism in delaying aging

[0063] Methods: The effects of active compounds on the metabolic phenotype of immune cells and Sirt1 protein expression were detected using a Seahorse cellular energy metabolism analyzer and Western blotting. The anti-aging effect of active compounds was evaluated in a classic model organism for aging studies—Saccharomyces cerevisiae cell model. Viable cell counts, Lüssler methylene blue staining, and qPCR were used to detect the effects of Polygonatum odoratum extract and active ingredients on yeast sequential aging and Sir2 gene expression.

[0064] Results: After intervention with rutin and methylophiopogonanone B, CD4 levels decreased. + T cell non-mitochondrial oxygen consumption increases, and intervention with Methylophiopogonanone B can also upregulate CD4. + T cell glycolysis levels and Sirt1 protein expression. In an in vitro yeast model (see detailed data below), it was found that Polygonatum odoratum extract (water extract), Rutin, and Methylophiopogonanone B could delay its senescence and increase Sir2 gene (a homolog of Sirt1) expression.

[0065] Specific data: Polygonatum odoratum extract and its active ingredients effectively delay yeast aging.

[0066] (a) Promoting yeast growth

[0067] Polygonatum odoratum extract inhibited yeast (BY4741) growth at a concentration of 100 mg / mL, and significantly promoted yeast growth at concentrations of 3.3 mg / mL and 0.37 mg / mL (P<0.01, P<0.05) (see...). Figure 10 ).

[0068] (II) Improving yeast sequential senescence

[0069] (1) Viable bacteria plate count

[0070] Depend on Figure 11 It was found that the survival rate of yeast cells gradually decreased with increasing culture time. The survival rates of the four groups of yeast cells did not change significantly between CLS 0 and CLS 3. After CLS 3, the survival rate of yeast cells began to decline rapidly, until the rate of decline slowed down at CLS 12. At CLS 6, the survival rate of yeast cells in the control group decreased to 58.3%, while the survival rate of 1 mg / mL Polygonatum odoratum extract was 86.9%, an increase of 49.05% compared to the control group. This indicates that Polygonatum odoratum extract can effectively delay yeast cell death, and the higher the concentration, the more significant the effect.

[0071] The effects of active compounds on the sequential senescence of yeast were detected by... Figure 12It was found that the survival rate of yeast cells gradually decreased with increasing culture time. The survival rates of the four groups of yeast cells did not change significantly between CLS 0 and CLS 4. After CLS 8, the survival rate of yeast cells began to decline rapidly, until the rate of decline slowed down at CLS 12. At CLS 12, the survival rate of yeast cells in the control group decreased to 46.8%, while the survival rates of 0.1 mg / mL Polygonatum odoratum extract, 0.01 mg / mL rutin, and 0.1 mg / mL methyl ophiopogon japonicus flavanone B were 79.5%, representing increases of 52.6%, 23.3%, and 69.9% respectively compared to the control group. This indicates that Polygonatum odoratum extract, rutin, and methyl ophiopogon japonicus flavanone B can effectively delay yeast cell death.

[0072] (2) Yeast staining observation

[0073] Lüsseau's basic methylene blue is a commonly used staining solution for assessing yeast cell viability. The staining agent, methylene blue, is blue in its oxidized state and colorless in its reduced state. When yeast cells are immersed in methylene blue solution, the stain enters the cells. Live cells, which produce reductase to convert methylene blue from its oxidized form to its reduced form, remain colorless, while dead cells, which cannot produce active reductase, are stained blue. Figure 13 It can be seen that the yeast cells in the blank senescent group began to show obvious dead cells, while the proportion of yeast cells stained blue in different doses of Polygonatum odoratum extract was smaller than that in the blank group, and the number of live yeast cells was significantly higher than that in the blank group. At the same time, it can be observed that the activity of Saccharomyces cerevisiae in different dose groups also showed significant differences. Yeast cells treated with 1-0.01 mg / mL had a higher proportion of live cells. The above results prove that Polygonatum odoratum extract has the effect of delaying the senescence of yeast cells.

[0074] Depend on Figure 14 It can be seen that the yeast cells in the blank senescent group began to show obvious dead cells, while the proportion of yeast cells stained blue in the rutin (0.1 mg / mL) and methyl ophiopogon flavanone B (0.1 mg / mL) groups was smaller than that in the blank group. That is, the yeast cells treated with rutin and methyl ophiopogon flavanone B had a higher proportion of live cells. The above results prove that rutin and methyl ophiopogon flavanone B have the effect of delaying the senescence of yeast cells.

[0075] (III) Upregulation of yeast Sir2 gene expression

[0076] Compared with the control group, the expression of the Sir2 gene in yeast in the model group was significantly decreased (P<0.001), and the expression of the Sir2 gene in yeast was significantly increased by Polygonatum odoratum extract at concentrations of 100, 10, 1, 0.1, and 0.01 mg / mL (P<0.05, P<0.01, P<0.001). Figure 15 ).

[0077] After yeast senescence, the effect of active ingredients from Polygonatum odoratum on the expression level of its Sir2 gene was investigated. Figure 16 The results showed that 1 mg / mL and 0.1 mg / mL of methyl ophiopogon flavanone B significantly increased the Sir2 gene content in senescent yeast (P<0.01).

[0078] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. Application of Polygonatum odoratum extract in the preparation of anti-aging drugs.

2. The application as described in claim 1, characterized in that, It is used to improve organ damage, age-related secretory phenotypes, and gut microbiota imbalance.

3. The application as described in claim 1 or 2, characterized in that, It is used to upregulate Sirt1 protein expression in the spleen and downregulate P16 protein expression in the spleen.

4. The application as described in claim 1, characterized in that, It is used to improve the composition of spleen immune cell subsets and aging-related metabolic phenotypes.

5. The application as described in claim 1 or 4, characterized in that, Used to significantly increase naive CD4 + T cell count, reduced memory (TEM) CD8 + The number of T cells was reduced, and the expression of P16 mRNA in spleen T cells was decreased.

6. The application as described in claim 1, characterized in that, Polygonatum odoratum extract improves CD4 by regulating the Park7 / Sirt1 signaling pathway + T-cell metabolism slows down aging.

7. The application as described in claim 1, characterized in that, It is used to delay yeast senescence and upregulate yeast Sir2 gene expression.

8. The application as described in any one of claims 1-7, characterized in that, The Solomon's Seal Extract is a water extract of Solomon's Seal.

9. The application as described in claim 8, characterized in that, The preparation method of the water extract of Polygonatum odoratum includes the following steps: take Polygonatum odoratum, add water and decoct 1-4 times, each time for 1-4 hours, centrifuge and filter the decoction, and concentrate the filtrate to obtain the product.

10. The application according to any one of claims 1 to 8, characterized in that, The Polygonatum odoratum extract, with the addition of excipients, is formulated into a clinically acceptable anti-aging pharmaceutical preparation.

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