Composite anti-aging nutritional capsule containing ergothioneine and nervonic acid and preparation method thereof

Abstract

According to the targeted anti-aging nutrition capsule and the preparation method thereof provided by the invention, the ergothioneine and the nervonic acid are formed by cooperatively regulating and controlling a PARP1-XRCC1-DNA ligase III complex, so that the DNA fracture repair efficiency is remarkably improved; a sodium hyaluronate-spermidine cationic liposome carrier is developed, and efficient co-delivery of water / fat double-soluble components is achieved; through clinical verification, triple anti-aging pathways of telomere maintenance, mitochondrial optimization and skin regeneration can be synchronously activated. The compound benefits of reducing DNA damage, improving skin elasticity and increasing NK cell activity can be realized.

Description

Technical Field

[0001] This invention relates to the field of functional foods and anti-aging preparations, specifically to a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid and its preparation method. Background Technology

[0002] Current anti-aging nutritional formulations face three major technological bottlenecks: First, mainstream products rely on single active ingredients, making it difficult to simultaneously activate multiple aging intervention pathways such as DNA repair, mitochondrial function, and skin regeneration; second, core ingredients such as nervonic acid and ergothioneine have low bioavailability due to differences in molecular polarity, and traditional carriers cannot achieve cross-barrier synergistic delivery; third, existing formulations are mostly limited to anti-oxidation or cellular energy replenishment, lacking the ability to regulate deep aging mechanisms such as telomere maintenance and targeted DNA damage repair. This invention utilizes a "dual-engine DNA repair + transmembrane delivery system" technical approach, leveraging the molecular complementarity of ergothioneine and nervonic acid, combined with cationic liposome carrier design, to achieve efficient absorption and targeted delivery of the ingredients. Simultaneously, it integrates mechanisms such as PQQ mitochondrial activation and spermine autophagy induction to construct a multi-dimensional anti-aging system, fundamentally reversing the cellular aging process. Summary of the Invention

[0003] The purpose of this section is to outline some aspects of embodiments of the present invention and to briefly describe some preferred embodiments. Simplifications or omissions may be made in this section, as well as in the abstract and title of this application, to avoid obscuring the purpose of these documents; however, such simplifications or omissions should not be construed as limiting the scope of the invention.

[0004] In view of the problems existing in the prior art, the inventors have proposed the present invention.

[0005] Therefore, the purpose of this invention is to overcome the shortcomings of the prior art and provide a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, and a preparation method thereof.

[0006] To solve the above-mentioned technical problems, the present invention provides the following technical solution: a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, comprising the following active components by weight percentage: ergothioneine 0.5-2%, disodium pyrroloquinoline quinone 1-3%, spermidine 0.8-2.5%, nervonic acid 15-25%, sodium hyaluronate 5-10%, type II collagen chondroitin powder 8-15%, Ganoderma lucidum extract 3-8%, lycopene 2-6%, ginseng extract 4-9%, black truffle extract 3-7%, taurine 1.5-4%, with the remainder being pharmaceutical excipients, wherein the nervonic acid is cis-15-tetracosenoyl nervonic acid.

[0007] As a preferred embodiment of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, wherein: the purity of nervonic acid is ≥95%, and its weight ratio with ergothioneine is (10-50):1; the β-glucan content in the Ganoderma lucidum extract is ≥30%, and it is prepared by supercritical CO2 extraction; the proportion of polypeptides with a molecular weight ≤5000Da in the black truffle extract is ≥60%.

[0008] As a preferred embodiment of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, the active components are encapsulated by cationic liposomes formed by sodium hyaluronate and spermidine, with a liposome particle size of 80-150 nm; the ergothioneine encapsulation rate in the liposomes is ≥90%, and the nervonic acid encapsulation rate is ≥92%.

[0009] A method for preparing an anti-aging nutritional capsule containing ergothioneine and nervonic acid complex includes the following steps: (a) Oil phase preparation: Nervonic acid and lycopene were dissolved in vegetable oil at 35-50℃ and homogenized; (b) Aqueous phase preparation: Sodium hyaluronate was dissolved in pure water, and disodium pyrroloquinoline quinone, ergothioneine, and spermidine were added and dispersed by ultrasonication; (c) Emulsification: The oil phase is injected into the aqueous phase and subjected to two-stage high-pressure homogenization to form a nanoemulsion; (d) Spray drying: Add a protective agent to the nanoemulsion and spray dry to obtain nanoparticles; (e) Solid-phase mixing: Type II collagen cartilage powder, Ganoderma lucidum extract, ginseng extract, black truffle extract, taurine and excipients are dry-mixed; (f) Capsule filling: Filling capsules with a mixture of nanoparticles and solid phase.

[0010] As a preferred embodiment of the preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, the two-stage high-pressure homogenization includes: a first-stage homogenization pressure of 40-60 MPa, 2-3 cycles; and a second-stage homogenization pressure of 120-150 MPa, 3-5 cycles. The inlet air temperature of the spray dryer is 170-190℃, the outlet air temperature is 75-85℃, the protective agent is maltodextrin, and the addition amount is 15-25% of the total solids.

[0011] As a preferred embodiment of the preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, wherein: the vegetable oil is coconut oil or medium-chain triglycerides; and the ultrasonic dispersion conditions are power 250-350W and time 10-20min.

[0012] As a preferred embodiment of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, it further comprises a DNA repair enhancer, wherein the DNA repair enhancer is selected from at least one of folic acid 0.1-0.5 mg / capsule or nicotinamide mononucleotide 10-30 mg / capsule.

[0013] As a preferred embodiment of the preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, wherein: when folic acid is added, its weight ratio with nervonic acid is 1:(600-800); when nicotinamide mononucleotide is added, its weight ratio with ergothioneine is (1-3):1.

[0014] As a preferred embodiment of the preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, the method further includes a quality control step: detecting the nervonic acid content using HPLC-ELSD method; and detecting the ergothioneine encapsulation rate using LC-MS / MS method.

[0015] As a preferred embodiment of the preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in this invention, the sample pretreatment for detecting the nervonic acid content includes: taking 0.1g of the capsule contents, adding n-hexane-isopropanol mixed solvent (1:1) and ultrasonically extracting for 30min.

[0016] The beneficial effects of this invention are as follows: Ergothioneine enhances cellular antioxidant capacity by activating the Nrf2 / ARE pathway, while simultaneously upregulating the efficiency of XRCC1-mediated DNA base excision repair; nervonic acid promotes neuronal membrane phospholipid remodeling and enhances PARP1 enzyme activity to accelerate DNA damage repair; PQQ and spermidine synergistically optimize mitochondrial function and activate mitophagy to clear senescent cells. This three-pathway linkage achieves comprehensive rejuvenation from the molecular to the cellular level.

[0017] This capsule utilizes a sodium hyaluronate-spermidine carrier to achieve highly efficient co-delivery of nervonic acid and ergothioneine. A two-stage high-pressure homogenization and spray drying process ensures that the average particle size of the nanoparticles is ≤150nm. Cellular experiments have verified that it can enhance DNA repair rates, and human trials have shown increased skin elasticity. Detailed Implementation

[0018] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.

[0019] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0020] Secondly, the term "an embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places throughout this specification does not necessarily refer to the same embodiment, nor is it a single embodiment or an embodiment selectively excluded from other embodiments.

[0021] Example 1 This embodiment provides a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, and its preparation method. Specifically, the basic formula preparation and delivery system construction are carried out according to the following steps: Raw material pretreatment Purification of nervonic acid: Take cis-15-tetracosene nervonic acid raw material (purity 92%), add 3 times its weight of ethanol and heat to 60℃ to dissolve. Preferably, remove free fatty acids by molecular distillation (vacuum degree 0.1mbar, temperature 130℃). The final product has an acid value ≤0.3mg KOH / g and a purity ≥98%. Preparation of Ganoderma lucidum extract: The fruiting bodies of Inonotus obliquus were pulverized to 40 mesh and extracted using supercritical CO2 (pressure 35 MPa, temperature 45℃). The β-glucan component was collected and freeze-dried, and the β-glucan content was found to be 38.7%. The Ganoderma lucidum extract was extracted using supercritical CO2 (pressure 35 MPa, temperature 45℃), and the β-glucan content was found to be 38.7%. Black truffle polypeptide extraction: Black truffle was degraded by a complex protease (papain: flavor protease = 1:2) at pH 6.0 and 50℃ for 6 hours, ultrafiltration was used to retain components with a molecular weight ≤5000Da, and freeze-drying was used to obtain active substances with a polypeptide content of 62.3%.

[0022] As one implementation method, an oil phase and an aqueous phase are prepared according to the following weight ratios: Oil phase: 180 mg nervonic acid and 30 mg lycopene dissolved in coconut oil (magnetically stirred at 45°C until completely dissolved); Aqueous phase: 60 mg of sodium hyaluronate was dissolved in 10 mL of 60 °C pure water, and 20 mg of pyrroloquinoline quinone disodium salt (PQQ), 15 mg of ergothioneine, and 18 mg of spermidine were added. The mixture was then further ultrasonically treated with a probe (300 W power, 15 min time) to disperse the molecules.

[0023] High-pressure homogenization and spray drying: The oil phase was injected into the aqueous phase, and two-stage pressure homogenization was performed: the first stage was 50 MPa for 2 cycles to break up large droplets, and the second stage was 150 MPa for 3 cycles to obtain nanoemulsions with a particle size D50=126nm (measured by Malvern laser particle size analyzer). Preferably, maltodextrin (20% of total solids) is added to the nanoemulsion as a protective agent, and the nanoparticles are prepared by spray drying in a spray drying tower (inlet air 185°C / outlet air 82°C, atomization pressure 0.3MPa), with an angle of repose of 28.3° and good flowability.

[0024] Solid-phase mixing and capsule filling: 100mg of type II collagen, 50mg of Ganoderma lucidum extract, 70mg of ginseng extract (ginsenoside Rg3 ≥ 5%), 40mg of black truffle extract, 25mg of taurine, and 392mg of microcrystalline cellulose were added to a V-type mixer and run at 15 rpm for 30 minutes. Finally, 5mg of magnesium stearate was added for lubrication, and the mixture was filled into size 0 capsules, each with a net weight of 800mg.

[0025] Quality control testing: Take 0.1g of capsule contents, add 10mL of n-hexane-isopropanol (1:1) mixed solvent, extract by ultrasonication for 30min, filter and inject the sample; Chromatographic conditions: C18 column (4.6×250mm, 5μm), mobile phase acetonitrile-isopropanol (85:15), flow rate 1.0mL / min, ELSD drift tube temperature 45℃; Ion source: ESI+, capillary voltage: 3.5kV, cone voltage: 30V, monitored ion pair m / z: 230.3→154.2.

[0026] Example 2 This embodiment provides a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, and its preparation method, specifically including delivery system optimization and cell-level anti-aging performance verification.

[0027] Feature Overview: The core innovation of sodium hyaluronate (HA)-spermidine cationic liposomes as a cross-barrier co-delivery system; Specifically, in pH 7.4 phosphate buffer, negatively charged HA (-35.2 mV) and positively charged spermidine (+28.7 mV) form a stable complex through charge neutralization.

[0028] An average particle size of 128±5nm (PDI=0.18) was achieved through two-stage high-pressure homogenization (first stage 50MPa demulsification → second stage 150MPa nano-sizing), breaking through the traditional limit of >200nm for emulsions (measured by Malvern laser particle size analyzer).

[0029] Alternative carrier solution: Chitosan-citric acid carrier (particle size 152nm, encapsulation efficiency 83.4%) Phospholipid bilayer carrier (DSPC:cholesterol = 2:1, encapsulation efficiency 89.1%) Preferably, 60 mg of HA and 18 mg of spermidine were dissolved in 10 mL of ultrapure water, magnetically stirred (600 rpm, 30 min), and allowed to stand for hydration for 2 h. After two-stage high-pressure homogenization, a milky white semi-transparent nanoemulsion was obtained, and the spherical structure was found to be intact under transmission electron microscopy.

[0030] Comparison of Encapsulation Efficiency and Stability (New Data Analysis) In-depth analysis of cellular-level anti-aging mechanisms DNA repair pathway validation Methods: Human dermal fibroblasts (HDF) were induced to damage by UVB with 0.5 J / cm², and then nanoparticles containing ergothioneine (10 μM) from Example 1 were added.

[0031] result: γH2AX focal number (DNA double-strand break marker): decreased by 78.9% in 24h (compared to 32.1% in the control group), demonstrating activation of the PARP1-XRCC1 repair complex.

[0032] Telomerase activity: 3.1-fold increase as detected by TRAP-PCR, and upregulation of key factors in the SIRT6 pathway by 2.4-fold (qPCR verification).

[0033] Quantitative evidence of collagen regeneration: Specifically, ELISA showed a 157% increase in type I collagen secretion and a 42% increase in the demethylation level of the COL1A1 gene promoter (methylation-specific PCR), confirming the epigenetic regulatory mechanism.

[0034] Justification for Enhanced Formulation Variants The technical necessity of folic acid supplementation: As a DNA methylation donor, folic acid (0.3 mg / capsule) promotes base excision repair (BER) of damaged DNA by providing methyl groups, and synergistically reduces homocysteine ​​levels with nervonic acid (a decrease of 38.2% as detected by HPLC-MS).

[0035] Bioequivalence of NMN as a substitute for ginseng extract: When the formula was replaced with NMN 20mg / capsule, the NAD+ level increased by 2.3 times (fluorescence method), and the SIRT6 pathway activation efficiency was not significantly different from the original formula (p>0.05).

[0036] Example 3 This embodiment provides a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, and its preparation method, specifically, a comprehensive efficacy evaluation of a randomized, double-blind clinical trial.

[0037] Subject stratification logic: Inclusion criteria: healthy women aged 50-65 (n=120), with skin elasticity R2 value ≤0.45 and serum telomere relative length ≤1.05 (qPCR method).

[0038] Exclusion criteria: Use of NAD+ precursors or DNA repair agents in the past 6 months.

[0039] Grouping method: Stratified randomization based on baseline telomere length (Block=4), the experimental group (60 people) took 2 capsules of Example 1 daily, and the control group (60 people) took an equal amount of starch capsules.

[0040] Standardization of evaluation indicators: Skin elasticity: R2 value (net elasticity / total deformation ratio) was measured using a Cutometer M580, under the same physician's operation and constant ambient temperature and humidity (25℃ / 50%RH).

[0041] DNA damage: single-cell gel electrophoresis (comet assay) was performed by double-blind pathologists to analyze tail moment values.

[0042] Multidimensional correlation analysis of results Cross-scale validation of typical cases Subject (62-year-old female): Skin biopsy: Masson staining showed a 46% increase in dermal collagen fiber density and a 3.2-fold upregulation of type IV collagen mRNA expression (in situ hybridization).

[0043] DNA repair kinetics: After UV-induced damage, the repair half-life was shortened from 4.2 h to 1.3 h (fitted by the comet tail moment decay model).

[0044] Molecular mechanism tracing: Mononuclear cell transcriptome sequencing showed that the enrichment of DNA repair pathway (KEGG: 03410) genes increased by 6.8 times (FDR<0.01).

[0045] Sufficientness of safety assessment Liver and kidney function monitoring: AST / ALT levels fluctuated by ≤15%, and creatinine clearance rate showed no significant change (p=0.37).

[0046] Adverse event record: 2 cases of mild gastrointestinal discomfort (experimental group), which were relieved after adjusting to take the medication after meals. There were no cases of withdrawal.

[0047] The three examples collectively demonstrate the core mechanism of "dual-engine DNA repair + mitochondrial optimization": At the molecular level: Ergothioneine increases the expression of XRCC1 repair protein by 3.8-fold by activating the Nrf2 / ARE pathway (thiolation modification of cysteine ​​residues in Keap1 protein); nervonic acid promotes neuronal membrane phospholipid remodeling (C24:1 ratio increases to 38.5%, as determined by LC-MS / MS), and enhances the binding efficiency of PARP1 to DNA break sites (SPR detection KD value increases from 10⁻). 7 M drops to 10⁻ 9 M).

[0048] At the cellular level: PQQ-spermidine increased mitochondrial membrane potential by 2.8 times (JC-1 red-green fluorescence ratio) and activated PINK1 / Parkin-mediated mitophagy (LC3-II / LC3-I ratio increased by 3.3 times).

[0049] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.

Claims

1. A compound anti-aging nutritional capsule containing ergothioneine and nervonic acid, characterized in that: The active ingredients include the following components by weight percentage: ergothioneine 0.5-2%, disodium pyrroloquinoline quinone 1-3%, spermidine 0.8-2.5%, nervonic acid 15-25%, sodium hyaluronate 5-10%, type II collagen chondroitin powder 8-15%, Ganoderma lucidum extract 3-8%, lycopene 2-6%, ginseng extract 4-9%, black truffle extract 3-7%, taurine 1.5-4%, with the balance being pharmaceutical excipients. The nervonic acid is cis-15-tetracosenoylnervonic acid.

2. The anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in claim 1, characterized in that: The purity of the nervonic acid is ≥95%, and its weight ratio with ergothioneine is (10-50):1; the β-glucan content in the Ganoderma lucidum extract is ≥30%, and it is prepared by supercritical CO2 extraction; the proportion of polypeptides with a molecular weight ≤5000 Da in the black truffle extract is ≥60%.

3. The anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in claim 2, characterized in that: The active component is encapsulated in cationic liposomes formed by sodium hyaluronate and spermidine, with a liposome particle size of 80-150 nm; the liposomes have an encapsulation rate of ergothioneine ≥90% and an encapsulation rate of nervonic acid ≥92%.

4. A method for preparing a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in any one of claims 1-3, characterized in that: Includes the following steps: (a) Oil phase preparation: Nervonic acid and lycopene were dissolved in vegetable oil at 35-50℃ and homogenized. (b) Aqueous phase preparation: Sodium hyaluronate was dissolved in pure water, and disodium pyrroloquinoline quinone, ergothioneine, and spermidine were added and dispersed by ultrasonication; (c) Emulsification: The oil phase is injected into the aqueous phase and subjected to two-stage high-pressure homogenization to form a nanoemulsion; (d) Spray drying: Add a protective agent to the nanoemulsion and spray dry to obtain nanoparticles; (e) Solid-phase mixing: Type II collagen cartilage powder, Ganoderma lucidum extract, ginseng extract, black truffle extract, taurine and excipients are dry-mixed; (f) Capsule filling: Filling capsules with a mixture of nanoparticles and solid phase.

5. The preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in claim 4, characterized in that: The two-stage high-pressure homogenization includes: a first-stage homogenization pressure of 40-60 MPa, with 2-3 cycles; and a second-stage homogenization pressure of 120-150 MPa, with 3-5 cycles. The inlet air temperature of the spray dryer is 170-190℃, the outlet air temperature is 75-85℃, the protective agent is maltodextrin, and the amount added is 15-25% of the total solids.

6. A method for preparing a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in any one of claims 4-5, characterized in that: The vegetable oil is coconut oil or medium-chain triglycerides; the ultrasonic dispersion conditions are power 250-350W and time 10-20min.

7. A compound anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in any one of claims 1-3, characterized in that: It also contains a DNA repair enhancer selected from at least one of folic acid 0.1-0.5 mg / capsule or nicotinamide mononucleotide 10-30 mg / capsule.

8. The anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in claim 7, characterized in that: When folic acid is added, its weight ratio to nervonic acid is 1:(600-800); when nicotinamide mononucleotide is added, its weight ratio to ergothioneine is (1-3):

1.

9. A method for preparing a compound anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in any one of claims 4-6, characterized in that: It also includes quality control steps: nervonic acid content is detected by HPLC-ELSD method; ergothioneine encapsulation rate is detected by LC-MS / MS method.

10. The preparation method of the anti-aging nutritional capsule containing ergothioneine and nervonic acid as described in claim 9, characterized in that: The sample pretreatment for the nervonic acid content detection includes: taking 0.1g of capsule contents, adding n-hexane-isopropanol mixed solvent (1:1), and ultrasonically extracting for 30min.

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