Anti-aging composition rich in PQQ and resveratrol and capable of activating longevity gene

Abstract

The invention provides an anti-aging composition rich in PQQ and resveratrol and capable of activating a longevity gene. The anti-aging composition comprises the following components in parts by mass: 5-50 parts of pyrroloquinoline quinone which is fermented and purified by microorganisms; 50 to 300 parts by mass of nano resveratrol; 50 to 500 parts by mass of coenzyme Q10 embedded by beta-cyclodextrin; 50-600 parts by mass of a taurine zinc complex; 40-150 parts by mass of an acanthopanax senticosus polysaccharide extract; 20-100 parts by mass of agrocybe cylindracea polysaccharide; 10 to 50 parts by mass of lycopene; 0.1-2 parts by mass of folic acid; and 50-500 parts by mass of betaine. According to the application, core components such as pyrroloquinoline quinone, resveratrol and coenzyme Q10 are integrated, and are matched with the taurine zinc complex and various plant active polysaccharides, so that a multi-target and multi-channel synergistic anti-aging system is constructed, and optimized microbial fermentation and nanocrystallization embedding processes are adopted, so that the purity and stability of key components are greatly improved.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine and functional foods, and particularly relates to an anti-aging composition rich in PQQ and resveratrol and activating longevity genes. Background Art

[0002] With the accelerating trend of global population aging, anti-aging research has evolved from a simple antioxidant theory to a multi-target and multi-pathway systematic intervention strategy. The essence of aging is the progressive decline of cell function over time, which involves multiple biological mechanisms such as mitochondrial dysfunction, decreased NAD+ levels, oxidative stress accumulation, and chronic inflammatory responses. Among them, mitochondria, as the energy factory of cells, the degradation of their function directly leads to insufficient ATP production and excessive free radicals, thereby triggering DNA damage and apoptosis. At the same time, the Sirtuin family of proteins, as NAD+-dependent deacetylases, has become an important target in anti-aging research by regulating metabolic balance, DNA repair, and inflammatory responses.

[0003] Among the existing anti-aging ingredients, pyrroloquinoline quinone (abbreviated as PQQ in English) and resveratrol show significant potential, but there are also obvious limitations. Although pyrroloquinoline quinone can promote mitochondrial biogenesis by activating PGC-1α and scavenge free radicals as a redox coenzyme, its bioavailability is low when used alone, and the oral absorption rate is usually less than 20%, and it is difficult to maintain a long-term antioxidant effect. Pyrroloquinoline quinone produced by chemical synthesis also has a risk of impurity residues, while the yield of the traditional fermentation method can only reach about 1000 μg / mL, and the production cost remains high. In addition, pyrroloquinoline quinone is sensitive to light and heat and requires complex embedding techniques to maintain stability, which greatly increases the formulation difficulty.

[0004] As a SIRT1 activator, resveratrol can inhibit the NF-κB pathway to exert anti-inflammatory and antioxidant effects, but its water solubility is extremely poor, with a solubility below 0.05 mg / mL at room temperature and an oral bioavailability of less than 1%. In the gastric acid environment, resveratrol is prone to degradation, and the yield of the traditional ethanol extraction method usually does not exceed 10%. The activity loss of unembedded resveratrol in simulated gastrointestinal fluid exceeds 60%. Although nano-carrier technologies such as ferritin embedding can improve stability, the encapsulation efficiency can only reach about 11.25%, and industrial production still faces major challenges.

[0005] Most existing anti-aging compositions adopt a simple stacking method, such as mechanically mixing NMN and resveratrol, lacking a systematic synergistic design. For example, although NMN can increase the NAD+ level, its effect depends on the activation of Sirtuins, while the rapid metabolism characteristic of resveratrol limits this synergistic effect. Due to its hydrophobicity, the bioavailability of coenzyme Q10 is less than 5%. Existing delivery systems such as liposomes not only have limited drug loading capacity but also poor storage stability.

[0006] From the perspective of clinical application, there are three main bottlenecks in current anti-aging technologies. First, there is insufficient multi-pathway synergy. Single components often only target specific aging pathways. For example, pyrroloquinoline quinone mainly acts on mitochondria, and resveratrol focuses on SIRT1 activation, making it difficult to comprehensively cover multi-dimensional aging mechanisms such as oxidative stress, inflammatory response, and metabolic disorders. Second, there are bottlenecks in component stability and delivery efficiency. Both pyrroloquinoline quinone and resveratrol require complex formulation processes such as nanonization or microencapsulation to improve stability, but existing technologies are difficult to achieve both high drug loading capacity and low-cost production simultaneously. Finally, there is a lack of long-term activation strategies. The activation of Sirtuins depends on the NAD+ level, and the half-life of NAD+ precursors such as NMN is only about 2 hours, requiring frequent dosing to maintain the effect.

[0007] Therefore, it is necessary to design an anti-aging composition rich in PQQ and resveratrol that activates longevity genes. Summary of the Invention

[0008] In order to overcome the defects in the existing technology, the present invention provides an anti-aging composition rich in PQQ and resveratrol that activates longevity genes.

[0009] To achieve the above object, the present invention provides the following technical solutions: An anti-aging composition rich in PQQ and resveratrol that activates longevity genes, calculated by mass, the composition comprises the following active components: Pyrroloquinoline quinone purified by microbial fermentation 5 - 50 parts by mass; Nanonized resveratrol 50 - 300 parts by mass; β-cyclodextrin-embedded coenzyme Q10 50 - 500 parts by mass; Taurine zinc complex 50 - 600 parts by mass; Acanthopanax senticosus polysaccharide extract 40 - 150 parts by mass; Agrocybe aegerita polysaccharide 20 - 100 parts by mass; Lycopene 10 - 50 parts by mass; Folic acid 0.1 - 2 parts by mass; Betaine 50 - 500 parts by mass.

[0010] The preparation method of the pyrroloquinoline quinone purified by microbial fermentation is as follows: a) Seed culture: Inoculate Hyphomicrobium denitrificans MQ004-2 into the seed medium, and shake-culture at 30-35 °C and 200-250 rpm for 20-24 hours until the OD600 reaches 1.5-2.0 to obtain a seed solution; b) Fermentation production: Transfer the seed solution into a fermenter at an inoculation amount of 5-10% wt, ferment at 35-37 °C for 60-72 hours, and supplement methanol every 6-8 hours to maintain the methanol concentration at 1.0-1.5 g / L to obtain a fermentation broth; c) Purification and crystallization: Filter the fermentation broth through a 0.22 μm microfiltration membrane, adsorb the filtrate with AB-8 macroporous resin, elute with 40-60% ethanol aqueous solution, concentrate the eluate to 1 / 5-1 / 3 of the original volume, add sodium chloride to a final concentration of 0.1-0.2 M, and stand and crystallize at 4-8 °C for 12-24 hours, and centrifuge and dry to obtain the pyrroloquinoline quinone purified by microbial fermentation.

[0011] The seed medium comprises the following components: containing 15-25 g / L of methanol, 2-3 g / L of ammonium sulfate, 5-6 g / L of disodium hydrogen phosphate, 0.5-1.5 g / L of magnesium sulfate heptahydrate, and the pH value of the seed medium is 6.5-7.5.

[0012] The fermentation medium comprises the following components: 20-30 g / L of methanol, 2-4 g / L of ammonium sulfate, 2-4 g / L of magnesium sulfate heptahydrate, control the dissolved oxygen at 20-30%, and the pH value of the fermentation medium is 6.8-7.2.

[0013] The preparation method of the nano-resveratrol comprises: a) Raw material extraction: Crush the rhizomes of Polygonum cuspidatum and mix them with 75% V / V ethanol at a material-liquid ratio of 1:10-20, and perform ultrasonic extraction at 60-70 °C for 2-3 times. Combine the extraction liquids and concentrate them under reduced pressure until there is no alcohol smell to obtain the crude resveratrol; b) Nano-emulsification: Dissolve the crude resveratrol and lecithin at a mass ratio of 1:1-3 in absolute ethanol, inject it into an aqueous phase containing 0.1-0.3% wt of poloxamer 188, and perform high-pressure homogenization at 12000-18000 psi for 4-6 times to obtain a nano-suspension with a particle size of 80-150 nm; c) Freeze-drying treatment: Add 5-10% wt of mannitol to the obtained nano-suspension as a freeze-drying protectant, pre-freeze at -50 °C to -80 °C for 4-6 hours, and perform vacuum freeze-drying for 24-36 hours to obtain the nano-resveratrol in the form of a white powder.

[0014] The power of the ultrasonic extraction is 200-400 W, the ultrasonic extraction frequency is 35-45 kHz, and each time is 20-40 minutes.

[0015] The preparation method of the β-cyclodextrin-embedded coenzyme Q10 comprises the following steps: a) Inclusion reaction: Dissolve coenzyme Q10 and β-cyclodextrin in an ethanol aqueous solution with a volume fraction of 70% V / V according to a molar ratio of 1:2 - 4, and stir and react at 50 - 60 °C for 3 - 5 hours; b) Spray drying: Spray dry the reaction solution to obtain a light yellow inclusion complex powder, which is the β-cyclodextrin-embedded coenzyme Q10, and the encapsulation efficiency of the β-cyclodextrin-embedded coenzyme Q10 is greater than 85%.

[0016] The inlet air temperature of the spray drying is 170 - 190 °C, and the outlet air temperature is 80 - 100 °C.

[0017] The preparation method of the zinc complex of taurine comprises: a) Coordination reaction: Mix taurine and zinc oxide according to a molar ratio of 1.8 - 2.2:1, add deionized water to a solid content of 15 - 25%, and stir and react at 75 - 85 °C for 3 - 5 hours to obtain a reaction solution; b) Purification and drying: Cool the reaction solution to 20 - 30 °C and then filter. Wash the filter cake with 70 - 90% ethanol 2 - 3 times, and vacuum dry at 55 - 65 °C for 10 - 14 hours to obtain a white product, which is the zinc complex of taurine.

[0018] The extraction method of the polysaccharide extract of Acanthopanax senticosus comprises: a) Water extraction and alcohol precipitation: Crush the root bark of Acanthopanax senticosus, add water according to a material-liquid ratio of 1:8 - 1:12, boil for 1 - 2 hours, concentrate the filtrate to 1 / 3 - 1 / 2 volume, and add 3 - 5 times the volume of 95% ethanol to precipitate the polysaccharide; b) Protein removal treatment: Dissolve the precipitate in water, and use the Sevage method to remove protein until there is no white interface, where the volume ratio of chloroform to n-butanol is 3 - 5:1; c) Freeze-dried product: Then dialyze for 48 - 72 hours and then freeze-dry to obtain a white powder with a polysaccharide content greater than 30%, which is the polysaccharide extract of Acanthopanax senticosus.

[0019] Compared with the prior art, the advantages and beneficial effects of the present invention are: 1. The anti-aging composition provided by this application realizes multi-dimensional intervention in the aging mechanism through the synergistic action of multiple components, and has significant advantages compared with the prior art. In terms of ingredient design, the composition not only contains two core anti-aging ingredients, pyrroloquinoline quinone and resveratrol, but also innovatively introduces auxiliary ingredients such as coenzyme Q10, zinc complex of taurine, and various plant polysaccharides, forming a complete action network covering mitochondrial function maintenance, oxidative stress regulation, inflammation inhibition, and metabolic balance regulation. This systematic design breaks through the limitation of single-target of traditional anti-aging products and can comprehensively intervene in the aging process at the molecular level.

[0020] 2. For pyrroloquinoline quinone, in this application, by optimizing the fermentation conditions of Microbacterium denitrificans, the methanol feeding concentration is precisely controlled at 1.0 - 1.5 grams per liter, and combined with the AB-8 macroporous resin purification process, the product purity is significantly improved. Aiming at the problem of poor solubility of resveratrol, the high-pressure homogenization method is used to prepare a nano-suspension with the particle size controlled in the range of 80 to 150 nanometers, and combined with poloxamer 188 as a stabilizer, greatly improving its absorption efficiency in the gastrointestinal tract. Coenzyme Q10 is encapsulated by β-cyclodextrin technology with an encapsulation rate of over 85%, solving the defect of insufficient drug loading capacity of traditional liposomes.

[0021] 3. Pyrroloquinoline quinone prepared by the microbial fermentation method has lower impurity content compared to the chemical synthesis method, and the addition of sodium chloride crystallization process enhances its thermal stability. Nano-resveratrol is freeze-dried and mannitol is added as a protective agent, which can maintain its activity at room temperature for a long time. The β-cyclodextrin encapsulation technology makes hydrophobic coenzyme Q10 form a water-soluble inclusion complex, avoiding the problem of oxidative degradation. These technological innovations together solve the industry problems of easy decomposition and difficult storage of anti-aging components.

[0022] 4. In the composition of this application, pyrroloquinoline quinone promotes mitochondrial biogenesis by activating PGC-1α, while coenzyme Q10 directly participates in the electron transport chain function, and the two jointly maintain mitochondrial energy metabolism. While nano-resveratrol activates the SIRT1 pathway, zinc-taurine complex provides zinc ions as a cofactor for Sirtuins, forming a cascade activation of NAD+-dependent deacetylase. Acanthopanax senticosus polysaccharide and Agrocybe aegerita polysaccharide indirectly affect the level of inflammatory factors by regulating the intestinal flora, complementing the NF-κB inhibitory effect of resveratrol. As a lipophilic antioxidant, lycopene and the aqueous phase free radical scavenging ability of zinc-taurine form a three-dimensional protection network, showing a unique synergistic effect among the components.

[0023] 5. In this application, a two-stage fermentation control strategy is adopted for pyrroloquinoline quinone. In the seed culture stage, the cell density is significantly increased by optimizing the ratio of ammonium sulfate and phosphate, and in the fermentation stage, the yield is increased by more than 30% through dynamic methanol feeding. The extraction of resveratrol innovatively uses the ultrasonic-assisted ethanol extraction method, operating at 60 to 70 degrees Celsius, which not only avoids high-temperature degradation but also improves the extraction efficiency. Acanthopanax senticosus polysaccharide is extracted by water extraction and alcohol precipitation combined with the Sevage method for protein deproteinization, and the polysaccharide yield is increased by about 20% compared to the traditional method. These process improvements have greatly reduced the production cost of each component, creating conditions for industrial production.

[0024] 6. In terms of clinical application value, the composition of the present application has multiple advantages. The synergistic effect of multiple active ingredients reduces the dosage requirement of a single ingredient and decreases the risk of side effects that may be brought about by high doses. Through the comprehensive application of nanonization and embedding technologies, the overall oral bioavailability is improved, reducing the dosing frequency. The introduction of zinc taurine not only provides zinc element, but also alleviates the liver metabolism burden through the hepatoprotective effect of taurine. Betaine, as a methyl donor, acts synergistically with folic acid to support the important epigenetic regulation mechanism of DNA methylation modification. Detailed implementation manners

[0025] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0026] In the present application, the models of various raw materials are briefly described as follows: All chemical reagents and raw materials comply with the Chinese Pharmacopoeia or food additive standards, and the equipment used in the production process is made of pharmaceutical-grade or food-grade materials.

[0027] Pyrroloquinoline quinone: Fermented and produced by the strain Hyphomicrobium denitrificans MQ004-2. The methanol used for fermentation is purchased from Sinopharm Chemical Reagent Co., Ltd., model GCS-500mL, purity ≥99.5%; ammonium sulfate is an Aladdin reagent, model A116166, analytical pure; disodium hydrogen phosphate is from Sigma-Aldrich, model S7907; magnesium sulfate heptahydrate is from Macklin Biochemical, model M813126.

[0028] Resveratrol: The raw material, rhizome of Polygonum cuspidatum, is purchased from Bozhou Chinese Medicinal Materials Market in Anhui, meeting the standards of the 2020 edition of the Chinese Pharmacopoeia. The ethanol used for extraction is produced by Sinopharm Chemical Reagent Co., Ltd., model 1000926831, 75% pharmaceutical grade. The nanonization auxiliary material lecithin is purchased from Shanghai Yuanye Bio-Technology Co., Ltd., model S24A9, soybean source purity ≥98%; poloxamer 188 is a product of BASF, model Kolliphor P188; the freeze-drying protectant mannitol is a Ron reagent, model R020235.

[0029] Coenzyme Q10: The raw material is purchased from Zhejiang Medicine Co., Ltd., model FQ1001, purity ≥99%; β-cyclodextrin is a product of Xi'an LvTian Biotechnology Co., Ltd., model CD-001, food grade; the ethanol used for embedding is from Sinopharm Chemical Reagent Co., Ltd., model 1000926831, 70% pharmaceutical grade.

[0030] Taurine zinc complex: Taurine is a product of Hubei Weideli Chemical Technology Co., Ltd., model WD-T1020, with pharmaceutical grade purity ≥99%; zinc oxide is a product of Beijing Bailingwei Technology Co., Ltd., model 901058, with nano-grade purity ≥99.9%; the ethanol used for purification is Aladdin reagent, model E111588, analytical grade.

[0031] Acanthopanax polysaccharide: Acanthopanax root bark was purchased from Jilin Fusong Ginseng Industry Co., Ltd. and meets the standards of the 2020 edition of the Chinese Pharmacopoeia. The ethanol used for precipitation was from Sinopharm Chemical Reagent Co., Ltd., model 1000926831, 95% industrial grade; the deproteinization reagent chloroform was from McLean Biochemical, model C805496; and n-butanol was from Aladdin Reagent, model B107172.

[0032] Agrocybe aegerita polysaccharide: Agrocybe aegerita fruiting bodies are purchased from the Gutian Edible Fungi Market in Fujian and comply with the GB 7096-2014 Edible Fungi Hygiene Standard.

[0033] Lycopene: The raw materials were purchased from Xinjiang Zhongji Natural Plant Purification Co., Ltd., model XJ-LYC-10, 10% natural tomato extract; the solvent soybean oil was a product of Cargill Grain and Oil Co., Ltd., model CJ-SO-100; the ethyl acetate used for extraction was from Sinopharm Chemical Reagent Co., Ltd., model 1000021011.

[0034] Folic acid: purchased from Jiangsu Jiangshan Pharmaceutical Co., Ltd., model JS-FA-100, pharmaceutical grade purity ≥99%, in line with USP standards.

[0035] Betaine: Purchased from Shandong Tianli Pharmaceutical Co., Ltd., model TL-BT-50, extracted from natural beets, purity ≥98%.

[0036] The technical solution of this application is: An anti-aging composition rich in PQQ and resveratrol and activating longevity genes, the composition comprising the following active components in parts by mass: 5-50 parts by weight of pyrroloquinoline quinone purified by microbial fermentation; 50-300 parts by weight of nano-resveratrol; 50-500 parts by weight of β-cyclodextrin-encapsulated coenzyme Q10; 50-600 parts by weight of taurine zinc complex; 40-150 parts by weight of Acanthopanax senticosus polysaccharide extract; 20-100 parts by weight of Agrocybe aegerita polysaccharide; 10-50 parts by weight of lycopene; Folic acid 0.1-2 parts by weight; Betaine 50-500 parts by mass.

[0037] The preparation method of the microbially fermented and purified pyrroloquinoline quinone is as follows: a) Seed culture: Inoculate Hyphomicrobium denitrificans MQ004-2 into the seed medium, and culture it by shaking at 30-35 °C and 200-250 rpm for 20-24 hours until the OD600 reaches 1.5-2.0 to obtain a seed solution; b) Fermentation production: Transfer the seed solution into a fermenter at an inoculation amount of 5-10% wt, and ferment at 35-37 °C for 60-72 hours. Methanol is supplemented every 6-8 hours to maintain the methanol concentration at 1.0-1.5 g / L to obtain a fermentation broth; c) Purification and crystallization: Filter the fermentation broth through a 0.22 μm microfiltration membrane, adsorb the filtrate with AB-8 macroporous resin, elute with 40-60% ethanol aqueous solution, concentrate the eluate to 1 / 5-1 / 3 of the original volume, add sodium chloride to a final concentration of 0.1-0.2 M, and let it stand and crystallize at 4-8 °C for 12-24 hours. Centrifuge and dry to obtain the microbially fermented and purified pyrroloquinoline quinone.

[0038] The seed medium comprises the following components: containing 15-25 g / L of methanol, 2-3 g / L of ammonium sulfate, 5-6 g / L of disodium hydrogen phosphate, 0.5-1.5 g / L of magnesium sulfate heptahydrate, and the pH value of the seed medium is 6.5-7.5.

[0039] The fermentation medium comprises the following components: 20-30 g / L of methanol, 2-4 g / L of ammonium sulfate, 2-4 g / L of magnesium sulfate heptahydrate, control the dissolved oxygen at 20-30%, and the pH value of the fermentation medium is 6.8-7.2.

[0040] The preparation method of the nano-resveratrol comprises: a) Raw material extraction: Crush the rhizome of Polygonum cuspidatum and mix it with 75% V / V ethanol at a solid-liquid ratio of 1:10-20, and perform ultrasonic extraction at 60-70 °C for 2-3 times. Combine the extraction solutions and concentrate them under reduced pressure until the alcohol smell disappears to obtain the crude resveratrol; b) Nano-emulsification: Dissolve the crude resveratrol and lecithin at a mass ratio of 1:1-3 in absolute ethanol, inject it into an aqueous phase containing 0.1-0.3% wt of poloxamer 188, and perform high-pressure homogenization at 12000-18000 psi for 4-6 times to obtain a nano-suspension with a particle size of 80-150 nm; c) Freeze-drying treatment: Add 5-10% wt of mannitol to the obtained nano-suspension as a freeze-drying protectant, pre-freeze at -50 °C to -80 °C for 4-6 hours, and perform vacuum freeze-drying for 24-36 hours to obtain the nano-resveratrol in the form of a white powder.

[0041] The power of the ultrasonic extraction is 200-400 W, the ultrasonic extraction frequency is 35-45 kHz, and each time is 20-40 minutes.

[0042] The preparation method of the β-cyclodextrin-embedded coenzyme Q10 comprises the following steps: a) Inclusion reaction: Dissolve coenzyme Q10 and β-cyclodextrin in an aqueous ethanol solution with a volume fraction of 70% V / V at a molar ratio of 1:2-4, and stir and react at 50-60 °C for 3-5 hours; b) Spray drying: Spray dry the reaction solution to obtain a light yellow inclusion complex powder, which is the β-cyclodextrin-embedded coenzyme Q10, and the encapsulation efficiency of the β-cyclodextrin-embedded coenzyme Q10 is greater than 85%.

[0043] The inlet air temperature of the spray drying is 170-190 °C, and the outlet air temperature is 80-100 °C.

[0044] The preparation method of the zinc taurine complex comprises: a) Coordination reaction: Mix taurine and zinc oxide at a molar ratio of 1.8-2.2:1, add deionized water until the solid content is 15-25%, and stir and react at 75-85 °C for 3-5 hours to obtain a reaction solution; b) Purification and drying: Cool the reaction solution to 20-30 °C and then filter, wash the filter cake with ethanol with a volume fraction of 70-90% for 2-3 times, and vacuum dry at 55-65 °C for 10-14 hours to obtain a white product, which is the zinc taurine complex.

[0045] The extraction method of the acanthopanax senticosus polysaccharide extract comprises: a) Water extraction and alcohol precipitation: Crush the acanthopanax senticosus root bark, add water according to a material-liquid ratio of 1:8-1:12, boil for 1-2 hours, concentrate the filtrate to 1 / 3-1 / 2 volume, and add 3-5 times the volume of 95% ethanol to precipitate the polysaccharide; b) Protein removal treatment: Dissolve the precipitate in water, and use the Sevage method to remove protein until there is no white interface, wherein the volume ratio of chloroform to n-butanol is 3-5:1; c) Freeze-dried product: Then dialyze for 48-72 hours and freeze-dry to obtain a white powder with a polysaccharide content greater than 30%, which is the acanthopanax senticosus polysaccharide extract.

[0046] In this application, by innovatively integrating core components such as pyrroloquinoline quinone purified by microbial fermentation, nano-sized resveratrol, and β-cyclodextrin-embedded coenzyme Q10, and combining with zinc taurine complex and various plant active polysaccharides, a multi-target and multi-pathway synergistic anti-aging system is constructed. Its significant advantages are reflected in the adoption of optimized microbial fermentation and nano-embedding processes, which greatly improve the purity and stability of key components; through the synergistic effect of four mechanisms including mitochondrial function enhancement, Sirtuins activation, antioxidant defense, and inflammation regulation, systematic intervention in the aging process is achieved; the innovative formula design enables significant synergistic effects among components, while reducing the dosage requirements of single components.

[0047] The technical solutions of the present invention are further illustrated below through examples and comparative examples, but the protection scope of the present invention is not limited thereto.

[0048] Example 1 The anti-aging composition provided in this example contains the following active components: 50 parts by mass of pyrroloquinoline quinone purified by microbial fermentation, 150 parts by mass of nano-resveratrol, 50 parts by mass of coenzyme Q10 embedded with β-cyclodextrin, 600 parts by mass of zinc taurine complex, 75 parts by mass of acanthopanax senticosus polysaccharide extract, 20 parts by mass of agrocybe aegerita polysaccharide, 50 parts by mass of lycopene, 1 part by mass of folic acid, and 500 parts by mass of betaine.

[0049] Preparation of pyrroloquinoline quinone: For seed culture, a medium containing 25 g / L of methanol, 3 g / L of ammonium sulfate, 6 g / L of disodium hydrogen phosphate, and 1.5 g / L of magnesium sulfate heptahydrate was used, with a pH of 7.5. Rhodobacter sphaeroides MQ004-2 was inoculated and cultured at 35 °C and 250 rpm for 24 hours until the OD600 reached 2.0. In the fermentation stage, it was transferred to a fermenter at an inoculation amount of 10%, fermented at 37 °C for 72 hours, and methanol was replenished every 8 hours to maintain a concentration of 1.5 g / L. During purification, AB-8 resin was eluted with 60% ethanol, sodium chloride was added to 0.2 M after concentration, and crystallization was carried out at 4 °C for 24 hours.

[0050] Preparation of nano-resveratrol: The rhizomes of polygonum cuspidatum and 75% ethanol were mixed at a solid-liquid ratio of 1:15, and ultrasonic extraction was carried out 3 times at 65 °C, with a power of 300 W and a frequency of 40 kHz, each time for 30 minutes. After the extract was concentrated, it was dissolved in ethanol with lecithin at a mass ratio of 1:2, injected into an aqueous phase containing 0.2% poloxamer 188, and homogenized at 15000 psi for 5 times. 7.5% mannitol was added during freeze-drying, and pre-freezing was carried out at -65 °C for 5 hours.

[0051] Embedding of coenzyme Q10: Coenzyme Q10 and β-cyclodextrin were dissolved in 70% ethanol at a molar ratio of 1:3, reacted at 55 °C for 4 hours, and spray-dried with an inlet air temperature of 180 °C and an outlet air temperature of 90 °C.

[0052] Zinc taurine complex: Taurine and zinc oxide were reacted at a molar ratio of 2:1, stirred at 80 °C for 4 hours, the filter cake was washed 3 times with 80% ethanol, and vacuum-dried at 60 °C for 12 hours.

[0053] Extraction of acanthopanax senticosus polysaccharide: The root bark of acanthopanax senticosus was extracted with water at a solid-liquid ratio of 1:10 for 1.5 hours, precipitated with 4 times the volume of 95% ethanol, deproteinized by the Sevage method, with a volume ratio of chloroform to n-butanol of 4:1, dialyzed for 60 hours and then freeze-dried.

[0054] Example 2 The composition of this example contains: 5 parts by mass of pyrroloquinoline quinone purified by microbial fermentation, 300 parts by mass of resveratrol, 500 parts by mass of coenzyme Q10, 50 parts by mass of zinc complex of taurine, 150 parts by mass of acanthopanax senticosus polysaccharide extract, 100 parts by mass of agrocybe aegerita polysaccharide, 10 parts by mass of lycopene, 2 parts by mass of folic acid, and 50 parts by mass of betaine.

[0055] Preparation of pyrroloquinoline quinone: The seed medium contains 15 g / L of methanol, 2 g / L of ammonium sulfate, 5 g / L of disodium hydrogen phosphate, 0.5 g / L of magnesium sulfate heptahydrate, pH 6.5, cultured at 30 °C and 200 rpm for 20 hours until OD600 reaches 1.5. In the fermentation stage, the methanol feeding concentration is 1.0 g / L, fermented at 35 °C for 60 hours, and eluted with 40% ethanol for purification, with the final concentration of sodium chloride being 0.1 M.

[0056] Nanometerization of resveratrol: The ultrasonic extraction power is 200 W, the frequency is 35 kHz, and each time is 20 minutes. The high-pressure homogenization pressure is 12000 psi, for 4 times, the freeze-drying protectant is 5% mannitol, and pre-frozen at -50 °C for 4 hours.

[0057] Embedding of coenzyme Q10: The molar ratio is 1:4, reacting at 60 °C for 5 hours, the inlet air temperature for spray drying is 190 °C, and the outlet air temperature is 100 °C.

[0058] Zinc complex of taurine: The molar ratio is 1.8:1, reacting at 75 °C for 3 hours, washing with 70% ethanol for 2 times, and drying at 55 °C for 10 hours.

[0059] Acanthopanax senticosus polysaccharide: The ratio of material to liquid is 1:12, the alcohol precipitation volume is 5 times, the Sevage method ratio is 5:1, and dialysis for 72 hours.

[0060] Example 3 The composition of this example contains: 25 parts by mass of pyrroloquinoline quinone purified by microbial fermentation, 50 parts by mass of resveratrol, 250 parts by mass of coenzyme Q10, 300 parts by mass of zinc complex of taurine, 40 parts by mass of acanthopanax senticosus polysaccharide extract, 60 parts by mass of agrocybe aegerita polysaccharide, 25 parts by mass of lycopene, 0.1 parts by mass of folic acid, and 250 parts by mass of betaine.

[0061] Preparation of pyrroloquinoline quinone: The seed medium contains 20 g / L of methanol, 2.5 g / L of ammonium sulfate, 5.5 g / L of disodium hydrogen phosphate, 1.0 g / L of magnesium sulfate heptahydrate, pH 7.0, cultured at 32 °C and 225 rpm for 22 hours until OD600 reaches 1.8. In the fermentation stage, the methanol feeding is 1.2 g / L, fermented at 36 °C for 66 hours, and eluted with 50% ethanol for purification, with the final concentration of sodium chloride being 0.15 M.

[0062] Nanization of resveratrol: ultrasonic power 400 W, frequency 45 kHz, 40 minutes each time. High-pressure homogenization at 18,000 psi, 6 times, freeze-drying protectant 10% mannitol, pre-freezing at -80 °C for 6 hours.

[0063] Coenzyme Q10 encapsulation: molar ratio 1:2, reaction at 50 °C for 3 hours, inlet air temperature of spray drying 170 °C, outlet air temperature 80 °C.

[0064] Zinc complex of taurine: molar ratio 2.2:1, reaction at 85 °C for 5 hours, washing with 90% ethanol 3 times, drying at 65 °C for 14 hours.

[0065] Acanthopanax senticosus polysaccharide: ratio of material to liquid 1:8, alcohol precipitation with 3 times the volume, Sevage method ratio 3:1, dialysis for 48 hours.

[0066] Comparative Example 1 In this comparative example, the same parts as in Example 1 will not be described in detail, and the differences are as follows: Zinc complex of taurine was not added, and the other components were the same.

[0067] Comparative Example 2 In this comparative example, the same parts as in Example 2 will not be described in detail, and the differences are as follows: Ordinary resveratrol powder was used instead of nanized resveratrol.

[0068] Comparative Example 3 In this comparative example, the same parts as in Example 3 will not be described in detail, and the differences are as follows: Coenzyme Q10 was not subjected to β-cyclodextrin encapsulation treatment.

[0069] Comparative Example 4 In this comparative example, the same parts as in Example 1 will not be described in detail, and the differences are as follows: The Acanthopanax senticosus polysaccharide extract was replaced with an equal amount of starch.

[0070] Comparative Example 5 In this comparative example, the same parts as in Example 2 will not be described in detail, and the differences are as follows: Pyrroloquinoline quinone was prepared by chemical synthesis method instead of microbial fermentation.

[0071] Test Results and Analysis The compositions of each example and comparative example were analyzed and tested according to relevant standards. By comparing the data of the examples and comparative examples, it can be seen that the technical solution of the present invention has significant advantages in many aspects, and the specific results are shown in Table 1.

[0072] Examples 1-3 were significantly superior to the comparative examples in terms of mitochondrial activity and SIRT1 activation levels, confirming the synergistic effect of pyrroloquinoline quinone and zinc taurine. The mitochondrial activity in Comparative Example 1 decreased by 16%, and the key value of nano-resveratrol was also demonstrated. The bioavailability in Comparative Example 2 decreased by 23%. β-cyclodextrin embedding improved the stability of coenzyme Q10, and the antioxidant capacity in Comparative Example 3 decreased by 7%. The purity of pyrroloquinoline quinone was increased by microbial fermentation method, and the mitochondrial activity in Comparative Example 5 decreased by 20%. Acanthopanax senticosus polysaccharide is irreplaceable for inflammation regulation, and the antioxidant capacity in Comparative Example 4 decreased by 16%.

[0073] Table 1 Test Results

[0074] This application achieves all-dimensional anti-aging through mitochondrial activation, including Sirtuins regulation of pyrroloquinoline quinone and coenzyme Q10, resveratrol and zinc, and the antioxidant network of lycopene and polysaccharide.

[0075] The above are the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle described in the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. An anti-aging composition rich in PQQ and resveratrol and activating longevity genes, characterized in that, The composition contains the following active components by mass parts: 5 - 50 mass parts of pyrroloquinoline quinone purified by microbial fermentation; 50 - 300 mass parts of nano-resveratrol; 50 - 500 mass parts of coenzyme Q10 embedded with β-cyclodextrin; 50 - 600 mass parts of zinc taurine complex; 40 - 150 mass parts of acanthopanax senticosus polysaccharide extract; 20 - 100 mass parts of agrocybe aegerita polysaccharide; 10 - 50 mass parts of lycopene; 0.1 - 2 mass parts of folic acid; 50 - 500 mass parts of betaine.

2. The anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 1, characterized in that, The preparation method of the pyrroloquinoline quinone purified by microbial fermentation is as follows: a) Seed culture: Inoculate Hyphomicrobium denitrificans MQ004-2 into the seed culture medium, and shake culture at 30 - 35 °C and 200 - 250 rpm for 20 - 24 hours until OD600 reaches 1.5 - 2.0 to obtain the seed liquid; b) Fermentation production: Transfer the seed liquid into the fermenter at an inoculation amount of 5 - 10%wt, ferment at 35 - 37 °C for 60 - 72 hours, and replenish methanol every 6 - 8 hours to maintain the methanol concentration at 1.0 - 1.5 g / L to obtain the fermentation broth; c) Purification and crystallization: Filter the fermentation broth through a 0.22 μm microfiltration membrane, adsorb the filtrate with AB-8 macroporous resin, elute with 40 - 60% ethanol aqueous solution, concentrate the eluate to 1 / 5 - 1 / 3 of the original volume, add sodium chloride to a final concentration of 0.1 - 0.2 M, stand and crystallize at 4 - 8 °C for 12 - 24 hours, and centrifuge and dry to obtain pyrroloquinoline quinone purified by microbial fermentation.

3. The anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 2, characterized in that, The seed culture medium includes the following components: containing 15 - 25 g / L of methanol, 2 - 3 g / L of ammonium sulfate, 5 - 6 g / L of disodium hydrogen phosphate, 0.5 - 1.5 g / L of magnesium sulfate heptahydrate, and the pH value of the seed culture medium is 6.5 - 7.

5.

4. The anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 2, characterized in that, The fermentation culture medium includes the following components: 20 - 30 g / L of methanol, 2 - 4 g / L of ammonium sulfate, 2 - 4 g / L of magnesium sulfate heptahydrate, control the dissolved oxygen at 20 - 30%, and the pH value of the fermentation culture medium is 6.8 - 7.

2.

5. An anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 1, characterized in that, The preparation method of the nano-resveratrol includes: a) Raw material extraction: Crush the rhizome of polygonum cuspidatum and mix it with 75% V / V ethanol at a material-liquid ratio of 1:10 - 20, perform ultrasonic extraction at 60 - 70 °C for 2 - 3 times, combine the extraction liquids and concentrate under reduced pressure until there is no alcohol smell to obtain the crude resveratrol; b) Nano-emulsification: Dissolve the crude resveratrol and lecithin at a mass ratio of 1:1 - 3 in absolute ethanol, inject it into the aqueous phase containing 0.1 - 0.3%wt of poloxamer 188, and perform high-pressure homogenization at 12000 - 18000 psi for 4 - 6 times to obtain a nano-suspension with a particle size of 80 - 150 nm; c) Freeze-drying treatment: Add 5 - 10%wt of mannitol to the obtained nano-suspension as a freeze-drying protectant, pre-freeze at -50 °C to -80 °C for 4 - 6 hours, and perform vacuum freeze-drying for 24 - 36 hours to obtain nano-resveratrol in the form of a white powder.

6. The anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 5, characterized in that: The power of the ultrasonic extraction is 200 - 400 W, the ultrasonic extraction frequency is 35 - 45 kHz, and each time is 20 - 40 minutes.

7. An anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 1, characterized in that The preparation method of the β-cyclodextrin-embedded coenzyme Q10 includes the following steps: a) Inclusion reaction: Dissolve coenzyme Q10 and β-cyclodextrin in an aqueous ethanol solution of 70% V / V at a molar ratio of 1:2 - 4, and stir and react at 50 - 60 °C for 3 - 5 hours; b) Spray drying: Spray dry the reaction solution to obtain a pale yellow inclusion complex powder, which is β-cyclodextrin-embedded coenzyme Q10. The encapsulation efficiency of the β-cyclodextrin-embedded coenzyme Q10 is greater than 85%.

8. An anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 7, characterized in that: The inlet air temperature of the spray drying is 170 - 190 °C, and the outlet air temperature is 80 - 100 °C.

9. An anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 1, characterized in that: The preparation method of the zinc taurine complex includes: a) Coordination reaction: Mix taurine and zinc oxide at a molar ratio of 1.8 - 2.2:1, add deionized water to a solid content of 15 - 25%, and stir and react at 75 - 85 °C for 3 - 5 hours to obtain a reaction solution; b) Purification and drying: Cool the reaction solution to 20 - 30 °C and then filter. Wash the filter cake with 70 - 90% ethanol 2 - 3 times, and vacuum dry at 55 - 65 °C for 10 - 14 hours to obtain a white product, which is the zinc taurine complex.

10. The anti-aging composition rich in PQQ and resveratrol and activating longevity genes according to claim 1, wherein: The extraction method of the Acanthopanax senticosus polysaccharide extract includes: a) Water extraction and alcohol precipitation: After crushing the root bark of Acanthopanax senticosus, add water according to a material-liquid ratio of 1:8 - 1:12 and boil for 1 - 2 hours. Concentrate the filtrate to 1 / 3 - 1 / 2 volume, and add 3 - 5 times the volume of 95% ethanol to precipitate the polysaccharide; b) Protein removal treatment: Dissolve the precipitate in water, and use the Sevage method to remove protein until there is no white interface, where the volume ratio of chloroform to n-butanol is 3 - 5:1; c) Freeze-dried product: Then dialyze for 48 - 72 hours and freeze-dry to obtain a white powder with a polysaccharide content greater than 30%, which is the Acanthopanax senticosus polysaccharide extract.

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