Application of quercetin-3-O-beta-D-glucuronide in preparation of anti-aging drugs
The anti-aging drug formulation prepared by quercetin-3-O-β-D-glucuronide solved the problem of lipofuscin accumulation in the Caenorhabditis elegans model, achieving the effects of prolonging life and delaying aging, demonstrating its application potential in anti-aging drugs and health products.
Patent Information
- Application Number
- CN202511892874.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-16
- Publication Date
- 2026-01-13
AI Technical Summary
In the existing technology, the anti-aging effect of quercetin-3-O-β-D-glucuronide has not been fully studied, and the Caenorhabditis elegans model has potential in observing the life-extending effect of drugs, but there is a lack of effective drug screening methods.
Quercetin-3-O-β-D-glucuronide was used as the active ingredient. It was prepared into various dosage forms such as injections, powders, granules, pills, oral liquids or tablets and applied to the Caenorhabditis elegans model. It significantly reduced lipofuscin accumulation and extended lifespan, suggesting its potential application in the preparation of anti-aging drugs and health products.
Quercetin-3-O-β-D-glucuronide significantly reduced the accumulation of lipofuscin in Caenorhabditis elegans and prolonged its lifespan, indicating that it has an anti-aging effect, and the effect is dose-dependent.
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Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the technical field of medicines, and provides a new application of quercetin-3-O-β-D-glucuronide, and particularly relates to an application of quercetin-3-O-β-D-glucuronide in preparation of an anti-aging medicine. BACKGROUND
[0002] Aging is caused by genetic and environmental factors and is an inevitable physiological change. With the passage of time, aging leads to some physiological phenomena, such as tissue protein degradation, tissue atrophy, and decreased metabolic rate, and can also lead to various diseases. Aging is one of the greatest risk factors for many chronic diseases, including cancer, cardiovascular and neurodegenerative diseases, which account for a large proportion of morbidity, mortality and medical costs worldwide (Zhang et al., 2024). According to the results of the eighth population census in 2022, China has officially entered a deep population aging stage. The China Statistical Yearbook 2024 shows that the population aged 65 and above in China has increased from 4.9% (in 1982) to 15.4% (in 2023), and the degree of aging is deepening. Various diseases caused by aging are increasingly burdening the social economy.
[0003] Many theories have been proposed about the mechanism of aging, one of which is that the excessive production of free radicals destroys the molecular and cellular structure, leading to the accumulation of the pro-aging factor malondialdehyde (MDA) in the body. MDA further damages cells to form lipofuscin, which is deposited in cells of various tissues and organs of the human body, leading to a slowdown in cell metabolism and a decrease in activity, thereby causing the functional decline of human organs, leading to aging. Therefore, lipofuscin is considered one of the important indicators of aging.
[0004] Caenorhabditis elegans is a small free-living animal that lives in temperate soil environments. As a classic model organism, it has been widely used in the study of obesity, aging, development, motor behavior, and neurodegenerative diseases. It has a short lifespan, low cost, easy cultivation, a strong nervous system, high conservation, and a large number of mutant strains. Data shows that 83% of the genes in the C. elegans proteome have homologous genes in humans, and 53% of the genes in the human coding genome can find homologous genes in C. elegans. Therefore, C. elegans has received widespread attention in the study of anti-aging (Link, 2006).
[0005] N2 is a wild-type strain of *C. elegans*. Due to its short lifespan, it can be used to observe the life-extending effects of drugs on *C. elegans* in the short term. Simultaneously, with age, lipofuscin accumulates in large quantities in the N2 gut as autofluorescent particles, exhibiting blue fluorescence. Because *C. elegans* is transparent, this blue fluorescence can be directly observed using a fluorescence microscope. The stronger the fluorescence intensity, the more severe the damage caused by free radicals in the nematode, and the more aged the nematode. Drugs with anti-aging effects can reduce the damage to cell structure caused by free radicals. After drug treatment, the weaker the blue fluorescence formed by lipofuscin in the nematode, the stronger the anti-aging effect of the drug.
[0006] Lotus leaf is a large, perennial aquatic herb, commonly used to treat bleeding, hyperlipidemia, obesity, AIDS, night sweats, fever, and inflammatory skin diseases. It is a typical plant used for both medicinal and culinary purposes. Lotus leaves are rich in flavonoids, with quercetin 3-O-β-D-glucuronide (Q3G) being particularly abundant. Q3G has been reported to exhibit various biological activities, such as antioxidant, anti-inflammatory, anti-atherosclerotic, and antiviral activities (Li et al., 2017). However, whether Q3G has anti-aging effects has not yet been studied.
[0007] Based on the above research status, this invention discloses a new application of quercetin-3-O-β-D-glucuronide. Quercetin-3-O-β-D-glucuronide can significantly reduce the accumulation of lipofuscin and prolong the lifespan of Caenorhabditis elegans, suggesting that quercetin-3-O-β-D-glucuronide has the potential to delay aging. It can be used in the preparation of anti-aging drugs and also in the preparation of health products that delay aging. Summary of the Invention
[0008] 1. The purpose of this invention is to provide an application of quercetin-3-O-β-D-glucuronide.
[0009] 2. Specifically, it involves the application of quercetin-3-O-β-D-glucuronide in the preparation of anti-aging drugs.
[0010] 3. The pharmaceutical preparation in which quercetin-3-O-β-D-glucuronide is the active ingredient is any dosage form such as injection, powder, granules, powder, pills, oral liquid, or tablets, which is understandable to those skilled in the art.
[0011] 4. The quercetin-3-O-β-D-glucuronide can also be used in the preparation of health products that delay aging.
[0012] 5. The dosage form of the health product containing quercetin-3-O-β-D-glucuronide as an active ingredient is any dosage form such as powder, granules, powder, pills, oral liquid, or tablets, which is understandable to those skilled in the art.
[0013] 6. The preparation method of quercetin-3-O-β-D-glucuronide according to the present invention is as follows:
[0014] Take 500 g of dried lotus leaves, mechanically pulverize them, and extract the pulverized medicinal material by heating and refluxing at 75°C with 95% ethanol three times (2 hours each time) to obtain the extract.
[0015] The extract was concentrated under reduced pressure using a rotary evaporator, and then further concentrated to a paste state by water bath evaporation. The weight was 26g.
[0016] Column chromatography was performed using D101 macroporous adsorption resin with elution gradients of water, 30% ethanol, 50% ethanol, 70% ethanol, and 100% ethanol. Thin-layer chromatography was used to detect similar components and combine them to obtain five fractions, numbered AE.
[0017] Part C was prepared by thin-layer chromatography to obtain a yellow amorphous powder, which is the quercetin-3-O-β-D-glucuronide.
[0018] 7. This invention uses a Caenorhabditis elegans model for experiments. The results show that quercetin-3-O-β-D-glucuronide can significantly reduce the accumulation of lipofuscin in Caenorhabditis elegans and prolong the lifespan of the nematodes. This suggests that quercetin-3-O-β-D-glucuronide has the potential to delay aging and can be used in the preparation of anti-aging drugs or health products that delay aging.
[0019] The technical solution of the present invention will be further described in detail below with reference to specific embodiments, but the scope of protection of the present invention is not limited to the following description. Attached Figure Description
[0020] Figure 1 The structural formula of quercetin-3-O-β-D-glucuronide.
[0021] Figure 2 The quercetin-3-O-β-D-glucuronide described in this invention delays the lifespan of N2 nematodes.
[0022] Figure 3 The quercetin-3-O-β-D-glucuronide described in this invention reduces the accumulation of lipofuscin in N2 nematodes. Detailed Implementation
[0023] Example 1
[0024] Preparation of quercetin-3-O-β-D-glucuronide according to the present invention:
[0025] Take 500 g of dried lotus leaves, mechanically pulverize them, and extract the pulverized medicinal material by heating and refluxing at 75°C with 95% ethanol three times (2 hours each time) to obtain the extract.
[0026] The extract was concentrated under reduced pressure using a rotary evaporator, and then further concentrated to a paste state by water bath evaporation. The weight was 26g.
[0027] Column chromatography was performed using D101 macroporous adsorption resin with elution gradients of water, 30% ethanol, 50% ethanol, 70% ethanol, and 100% ethanol. Thin-layer chromatography was used to detect similar components and combine them to obtain five fractions, numbered AE.
[0028] Part C was prepared by thin-layer chromatography to obtain a yellow amorphous powder, which is the quercetin-3-O-β-D-glucuronide.
[0029] Example 2
[0030] The life-prolonging effect of quercetin-3-O-β-D-glucuronide described in this invention on Caenorhabditis elegans N2:
[0031] 1. Biomaterials
[0032] (1) *C. elegans* N2 strain was purchased from the Caenorhabditis Genetics Center (CGC) and is a wild-type strain. N2 is a wild-type strain of *C. elegans*. Due to its short lifespan, it can be used to observe the life-extending effect of drugs on *C. elegans* in the short term. Simultaneously, with age, lipofuscin accumulates in large quantities in the N2 gut in the form of autofluorescent particles, exhibiting blue fluorescence. Since *C. elegans* is transparent, this blue fluorescence can be directly observed using a fluorescence microscope. The stronger the fluorescence intensity, the more severe the damage caused by free radicals in the nematode, and the more aged the nematode. Drugs with anti-aging effects can reduce the damage to cell structure caused by free radicals. Therefore, the longer the lifespan of the nematode and the weaker the intensity of the lipofuscin blue fluorescence, the stronger the anti-aging effect of the drug. In this embodiment, the *C. elegans* N2 strain was used as a model for screening anti-aging drugs.
[0033] (2) Escherichia coli OP50 (uracil leak mutant), purchased from Caenorhabditis Genetics Center (CGC), was used as food for Caenorhabditis elegans.
[0034] 2. Reagents
[0035] (1) Composition and preparation of solid NGM (Nematode Growth Medium) culture medium (taking 100 ml as an example): Ingredients Amount NaCl 0.3031 g Proteose peptone 0.2719 g KH2PO4 1.7033 g K2HPO4 0.2309 g Agar 1.7155 g Distilled water 100 mL
[0036] After preparing the solid NGM medium, autoclave it at 121°C for 20 min. Under aseptic conditions, add 100 µL of 5 mg / mL cholesterol, 100 µL of 1 M MgSO4, 100 µL of 1 M CaCl2, and 100 µL of 50 mM 5-fluorouracil. Shake well and pour the mixture into sterilized 6 cm culture plates while still hot, approximately 15 mL per plate. Let the medium solidify and set aside.
[0037] (2) M9 solution formula (taking 100 ml as an example): Ingredients Amount NaCl 0.4972 g KH2PO4 0.2992 g Na2HPO4 0.5964 g MgSO4 0.012 g Distilled water 100 mL
[0038] (3) Preparation of lysis buffer: 6.4% NaClO solution and 1M NaOH solution are mixed at a volume ratio of 1:1.
[0039] 3. Preparation of NGM plates containing quercetin-3-O-β-D-glucuronide
[0040] Weigh 19.12 mg of quercetin-3-O-β-D-glucuronide and dissolve it in 500 µL of DMSO to prepare a stock solution with a concentration of 80 mM. Add NGM medium to final concentrations of 20 µM, 40 µM, and 80 µM, respectively. Pour the NGM solution into each agar plate and allow it to solidify. Spread Escherichia coli OP50 evenly on the medium as food for the nematodes.
[0041] 4. Implementation Steps
[0042] (1) Cultivation of nematodes:
[0043] Caenorhabditis elegans was inoculated onto solid NGM plates coated with Escherichia coli OP50 and then cultured in an incubator at 15°C. When the nematodes grew to adulthood, they were synchronized.
[0044] (2) Nematode synchronization:
[0045] When the NGM medium contains a large number of adult nematodes and some eggs have hatched, flush the nematodes off the medium with M9 buffer and transfer them to a 5 mL centrifuge tube. Let the nematodes settle to the bottom of the tube and discard the supernatant. Add 2 mL of lysis buffer to the centrifuge tube and vortex for 5-7 minutes until all the nematodes break down. Stop vortexing and aliquot the mixture into 1.5 mL centrifuge tubes. Wash the nematode eggs three times with M9 solution.
[0046] (3) Experimental group setup
[0047] Blank control group: DMSO;
[0048] Positive control group: 100µM (resveratrol);
[0049] Quercetin-3-O-β-D-glucuronide group: 20 µM; 40 µM; 80 µM.
[0050] (4) Experimental steps
[0051] Synchronized nematode eggs were aliquoted into centrifuge tubes and incubated in M9 buffer for 30 hours to develop synchronized L1-stage nematode larvae. 10 µL of each larvae was counted three times, and the average value was taken. The nematodes were then transferred to NGM medium containing different concentrations of quercetin-3-O-β-D-glucuronide. The blank control group was NGM medium coated with OP50 and containing an equal volume of DMSO as the quercetin-3-O-β-D-glucuronide group. Resveratrol was used as the positive control. Eighty nematodes were inoculated into each culture dish, with three replicates as independent parallels. The nematodes were cultured at 20°C, and the number of dead nematodes was recorded daily until all nematodes died. For the nematode group used to measure lipofuscin blue fluorescence, the nematodes were washed off with M9 buffer on day five, centrifuged to remove the supernatant, and then anesthetized with 25 mM NaN3. The nematodes were observed and photographed under a fluorescence microscope. Forty nematodes were randomly selected from each group, and the fluorescence intensity of all images was quantitatively analyzed using ImageJ software.
[0052] (5) Experimental Results
[0053] Figure 2 The vertical axis represents the survival rate; a higher survival rate indicates a stronger life-extending effect of quercetin-3-O-β-D-glucuronide. Figure 3 The ordinate represents the fluorescence intensity of lipofuscin. The weaker the fluorescence intensity, the less damage caused by free radicals in the nematode, and the stronger the anti-aging effect of the drug. **** indicates p < 0.0001.
[0054] Experimental results showed that, in this embodiment, quercetin-3-O-β-D-glucuronide significantly reduced the accumulation of lipofuscin in *C. elegans* and prolonged the lifespan of the nematodes, suggesting that quercetin-3-O-β-D-glucuronide has the effect of prolonging lifespan and delaying aging. Furthermore, the drug effect was enhanced with increasing drug concentration, indicating that the anti-aging effect of quercetin-3-O-β-D-glucuronide is dose-dependent.
[0055] The above examples demonstrate that quercetin-3-O-β-D-glucuronide can significantly reduce the accumulation of lipofuscin in *C. elegans* and prolong its lifespan, suggesting that quercetin-3-O-β-D-glucuronide has the potential to delay aging and can be used in the preparation of anti-aging drugs or health products that delay aging. References [1] Zhang KZ et al. Petunidin-3-O-[rhamnopyranosyl-(trans-p-coumaroyl)]-5-O(β-D-glucopyranoside), the main anthocyanin from the fruits of Lyciumruthenicum murray, enhances the lifespan of Caenorhabditis elegans byactivating DAF-16 and improving the gut microbiota. "Food Bioscience", 2024, Volume 64, Page 104642. [2]Link. C. elegans models of age-associated neurodegenerativediseases: Lessons from transgenic worm models of Alzheimer's disease. "Experimental Gerontology", 2006, Volume 41, Pages 1007-1013. [3] Li F et al. Enrichment and separation of quercetin-3-O-β-d-glucuronide from lotus leaves (nelumbo nucifera gaertn.) and evaluation of its anti-inflammatory effect. "Journal of chromatography. B", 2016, Volume 1040, Pages 186-191.
Claims
1. The application of quercetin-3-O-β-D-glucuronide in the preparation of anti-aging drugs, wherein the chemical structure of quercetin-3-O-β-D-glucuronide is as follows: 。 2. The application as described in claim 1, characterized in that, The pharmaceutical preparations in which quercetin-3-O-β-D-glucuronide is the active ingredient include injections, powders, granules, pills, oral liquids, and tablets.
3. The application as described in claim 1, characterized in that, The preparation method of the quercetin-3-O-β-D-glucuronide is as follows: Take 500 g of dried lotus leaves, mechanically pulverize them, and extract the pulverized medicinal material by heating and refluxing at 75℃ with 95% ethanol three times (2 h each time) to obtain the extract. Concentrate the extract under reduced pressure using a rotary evaporator, and then continue to concentrate it into a paste by evaporation in a water bath, weighing 26 g. Perform column chromatography using D101 macroporous adsorption resin, eluting with water, 30% ethanol, 50% ethanol, 70% ethanol, and 100% ethanol as gradients. Through thin-layer chromatography detection, similar components were combined, and finally 5 fractions were obtained, numbered AE. Fraction C was obtained as a yellow amorphous powder by preparative thin-layer chromatography, which is the quercetin-3-O-glucuronide.