A multi-channel composite nutritional preparation for improving vascular endothelial function and a preparation method thereof

Abstract

This invention discloses a compound nutritional preparation for synergistically improving vascular endothelial function through multiple pathways and its preparation method. The preparation comprises L-citrulline, L-arginine, magnesium salt, vitamin K2, nattokinase, curcumin, naringin, vitamin C, vitamin E, betaine, γ-aminobutyric acid, black pepper powder, and excipients. The weight ratio of L-citrulline to L-arginine is 3:1-4:1, and the weight ratio of curcumin to naringin is 3:1-8:1. This invention is the first to construct a four-dimensional synergistic system of "NO pathway activation + vascular calcification inhibition + fibrinolytic activity maintenance + anti-inflammatory and antioxidant network". This invention uses liposome inclusion technology to improve the bioavailability of curcumin and naringin (AUC increased by 8.96 times, Cmax increased by 9.19 times), and uses enteric-coated microparticle technology to protect nattokinase activity (gastric juice activity retention rate increased from 23% to 94%). Experiments show that the formulation of this invention can reduce the cardio-ankle vascular index (CAVI) by 16.8% and high-sensitivity C-reactive protein (hs-CRP) by 67.8%, significantly outperforming the closest existing technology. Decomposed formulation experiments demonstrate a non-linear synergistic effect among the various modules. This invention improves vascular endothelial function through multi-pathway synergistic effects and is suitable for vascular function management in high-risk groups for arteriosclerosis and sub-healthy individuals.

Description

Technical Field

[0001] This invention belongs to the field of special dietary food technology, specifically relating to a compound nutritional preparation that synergistically improves vascular endothelial function through multiple pathways and its preparation method. This invention is particularly suitable for high-risk groups for arteriosclerosis, individuals at risk of cardiovascular diseases such as hypertension, hyperlipidemia, and diabetes, as well as sub-healthy individuals seeking proactive vascular health management. Background Technology

[0002] Cardiovascular disease (CVD) is the leading cause of death worldwide. According to the "China Cardiovascular Health and Disease Report 2022," the number of people with cardiovascular disease in my country is estimated to be as high as 330 million, and this number continues to rise. While existing standard drug treatments (such as statins, ACEI / ARBs, and biguanides) can effectively lower low-density lipoprotein cholesterol (LDL-C) and blood pressure, meta-analyses of large-scale clinical trials have shown that even if LDL-C is reduced to <1.8 mmol / L and blood pressure to <130 / 80 mmHg, the residual relative risk of cardiovascular events remains as high as 30-50% [Reference: Libby P, et al. Circulation. 2019;139(25):2897-2909]. This "residual risk paradox" suggests that simply intervening in traditional risk factors cannot fully restore the integrity of vascular endothelial function. Endothelial dysfunction is the initiating factor of atherosclerosis and the core pathological basis for the occurrence and development of cardiovascular events. Therefore, the new paradigm of CVD, from "data control" to "managing vascular health", has become a consensus among international authoritative institutions and experts.

[0003] In recent years, various compositions for improving vascular endothelial function have been reported. A search revealed that the prior art closest to this invention includes: Prior art document 1: CN2023111XXXXA (publication date September 2023), entitled "A nutritional complex for delaying vascular aging and its preparation method and application", applicant: Wuhan Huana United Pharmaceutical Co., Ltd. This patent discloses a nutritional complex comprising whey protein powder, maltodextrin, celery extract, bayberry extract, green tea extract, buckwheat extract, and multivitamins. The difference between prior art document 1 and this invention is that prior art document 1 relies on the mixed effect of multiple plant extracts and does not explicitly target the NO pathway or vascular calcification; its core components do not contain citrulline, arginine, vitamin K2, nattokinase, or naringin; nor does it employ liposome or enteric-coated formulation technology.

[0004] Prior art document 2: CN201910687695.8 (publication date 2021), entitled "A composition for reducing vascular endothelial damage in hypertensive patients," applicant: Shenzhen Aosa Pharmaceutical Co., Ltd. This patent discloses a composition containing 150-500 mg of curcumin and 0.5-100 mg of B vitamins. The difference between prior art document 2 and this invention is that prior art document 2 is a simple combination of "curcumin + B vitamins," while this invention uses an anti-inflammatory and antioxidant network of "curcumin + naringin + VC / VE"; prior art document 2 does not include the citrulline / arginine NO pathway module, the vitamin K2+ magnesium calcification inhibition module, or the nattokinase fibrinolytic module.

[0005] Prior art document 3: CN201980042798.7 (publication date 2021), entitled "Composition for Improving Vascular Endothelial Function or Peripheral Vascular Blood Flow", applicant: House Foods Health Products Co., Ltd. (Japan). This patent discloses a composition with curcuminol A, curcuminol B, and bisabolol as active ingredients. The difference between this invention and prior art document 3 is that prior art document 3 only relates to vasodilation and does not involve vascular calcification inhibition, fibrinolytic balance, or low-grade inflammation suppression.

[0006] Prior art document 4: CN200780013375.X (granted in 2013), entitled "Arteriosclerosis Prevention Agent, Inhibitor of Vascular Intimal Thickening, and Improver of Vascular Endothelial Function," applicant: Carpis Corporation (Japan). This patent discloses a composition containing Xaa Pro Pro tripeptide or Lactobacillus helveticus ferment. The difference between this invention and the present invention is that the active ingredient in prior art document 4 is a milk-derived tripeptide / fermentation product, while the present invention is a system integration of amino acids + plant extracts + enzymes + vitamins / minerals.

[0007] Prior art document 5: CN201710999148.4 (publication date 2018), entitled "A pharmaceutical composition for improving vascular endothelial dysfunction and its preparation method and use," applicant: Fujian University of Traditional Chinese Medicine. This patent discloses a traditional Chinese medicine compound composed of Gastrodia elata, Uncaria rhynchophylla, Nelumbo nucifera seed heart, and Scutellaria baicalensis. The difference between prior art document 5 and this invention is that prior art document 5 is a traditional Chinese medicine compound with complex and unclear active ingredients; this invention uses a clearly defined single compound component, and the dosage of each component is precisely controllable.

[0008] In summary, existing technologies share the following common drawbacks: First, the targets are singular. The aforementioned comparative documents either focus on the single activation of the NO pathway, or on antioxidation, or on the multi-target effects of crude extracts of traditional Chinese medicine but with unclear mechanisms. None of them simultaneously address the four core issues of "vasodilation, inhibition of vascular calcification, maintenance of fibrinolytic balance, and suppression of low-grade inflammation".

[0009] Second, there is a lack of evidence for synergistic compatibility among multiple modules. Existing technologies have not revealed the effect of a specific citrulline / arginine ratio on sustained NO production, nor have they revealed the synergistic effect of curcumin and naringin in inhibiting the NF-κB and NLRP3 inflammatory pathways.

[0010] Third, low bioavailability. The active ingredients, such as curcumin and naringin, have extremely poor water solubility, with oral bioavailability of less than 5%; nattokinase is rapidly inactivated in the acidic environment of the stomach. Existing patents have not employed liposome encapsulation or enteric coating technologies to address these issues.

[0011] Therefore, developing a compound nutritional preparation with multi-pathway synergy, high bioavailability, and precisely optimized dosage of each component has significant clinical and industrial value. Summary of the Invention

[0012] I. Summary of the Invention This invention provides a compound nutritional preparation that synergistically improves vascular endothelial function through multiple pathways. The distinguishing technical features of this invention compared to existing technologies are as follows: (1) For the first time, L-citrulline and L-arginine were combined in a specific weight ratio (3.2:1 to 3.8:1) to achieve continuous NO generation and avoid the first-pass metabolism problem of liver when directly supplementing arginine; (2) For the first time, vitamin K2 and magnesium salts were combined for the inhibition of vascular calcification, and formed a complete functional closed loop with the NO pathway module; (3) For the first time, naringin and curcumin were combined in a specific weight ratio (4:1 to 6:1) and treated with liposome inclusion technology to achieve dual inhibition of the NF-κB and NLRP3 inflammatory pathways, resulting in a synergistic effect. (4) For the first time, enteric-coated microparticle technology was used to protect the activity of nattokinase, and it was prepared as a single formulation together with the above modules; (5) For the first time, four functional modules, namely “NO pathway activation, vascular calcification inhibition, fibrinolytic activity maintenance, and anti-inflammatory and antioxidant network”, are integrated into one formulation to achieve systematic repair of vascular endothelial function.

[0013] II. Technical Solution This invention provides a compound nutritional preparation for synergistically improving vascular endothelial function through multiple pathways, characterized in that it is composed of the following raw materials in parts by weight and is divided into three protective layers: (a) Wide-range protection scheme: 60-80 parts L-citrulline, 15-25 parts L-arginine, 5-12 parts magnesium salt, 0.8-1.8 parts vitamin K2, 10-20 parts nattokinase, 3-8 parts curcumin, 0.5-2 parts naringin, 3-8 parts vitamin C, 0.1-0.5 parts vitamin E, 12-22 parts betaine, 1-4 parts γ-aminobutyric acid, and 0.2-0.6 parts black pepper powder.

[0014] (II) Medium-range protection scheme: 65-75 parts L-citrulline, 18-22 parts L-arginine, 6-10 parts magnesium salt, 1.0-1.5 parts vitamin K2, 14-18 parts nattokinase, 4-6 parts curcumin, 0.8-1.2 parts naringin, 4-6 parts vitamin C, 0.15-0.3 parts vitamin E, 14-18 parts betaine, 2-3 parts γ-aminobutyric acid, and 0.3-0.5 parts black pepper powder.

[0015] (III) Narrow-range protection scheme (best implementation method): 70 parts L-citrulline, 20 parts L-arginine, 25 parts phosphatidylcholine, 8 parts magnesium gluconate, 1.2 parts vitamin K2 (MK-7), 16 parts nattokinase, 5 parts curcumin, 1 part naringin, 4.5 parts vitamin C, 0.2 parts vitamin E, 16 parts betaine, 2.4 parts γ-aminobutyric acid, 0.37 parts black pepper powder, 68 parts isomaltooligosaccharide, 61.33 parts maltodextrin, 0.8 parts neotame, and 0.2 parts pineapple flavoring.

[0016] Key feature constraints: The weight ratio of L-citrulline to L-arginine is 3:1 to 4:1, preferably 3.2:1 to 3.8:1, and most preferably 3.5:1.

[0017] The weight ratio of curcumin to naringin is 3:1 to 8:1, preferably 4:1 to 6:1, and most preferably 5:1.

[0018] III. Formulation Technology (1) Curcumin-naringin liposome inclusion complex Formula: Curcumin: Naringin: Soy phosphatidylcholine = 1:0.2:5 (by weight) Preparation method: Dissolve the above components in anhydrous ethanol, evaporate at 45°C to form a film, hydrate with pH 7.4 phosphate buffer, and homogenize by high pressure microfluidic jet (800 bar, 5 cycles). Technical parameters: Average particle size 80-120nm, encapsulation efficiency ≥85% (2) Nattokinase enteric-coated microparticles Coating material: Eudragit L100-55 (methacrylic acid copolymer) Drug-loaded pellet core: microcrystalline cellulose Preparation method: Coating by bottom spraying in a fluidized bed, inlet air temperature 42±2℃, spraying rate 1.5ml / min Release characteristics: <5% release rate in 0.1M hydrochloric acid after 2 hours; >85% release rate in pH 6.8 phosphate buffer after 30 minutes. (3) Vitamin K2 microcrystal co-treatment product Process: Vitamin K2 and microcrystalline cellulose are spray-dried together at a weight ratio of 1:10. Stability: After being stored at 40℃ and 75%RH for 6 months, the content decreased by <5%.

[0019] IV. Preparation Method (1) Preparation of liposomes: Curcumin, naringin and phosphatidylcholine were dissolved in anhydrous ethanol at a weight ratio of 1:0.2:5, and the mixture was evaporated at 45°C to form a film. The film was hydrated by adding pH 7.4 phosphate buffer, cyclically heated 5 times at 800 bar using a high-pressure microfluidic homogenizer, passed through a 0.22 μm membrane, and spray-dried to obtain liposome powder. (2) Preparation of enteric-coated microcapsules: Nattokinase and Eudragit L100-55 were suspended in 60% ethanol at a weight ratio of 1:2. Microcrystalline cellulose pellet cores were used as substrates and coated by bottom spraying in a fluidized bed. The air inlet temperature was 42±2℃ and the spraying rate was 1.5ml / min. After granulation, the pellets were dried at 40℃ for 2 hours. (3) Preparation of direct-mixed powder: L-citrulline, L-arginine, magnesium gluconate, vitamin K2 microcrystal co-treated product, betaine, GABA, vitamin C, vitamin E, black pepper powder, isomaltooligosaccharide, maltodextrin, neotame, and pineapple flavoring are put into a three-dimensional motion mixer and mixed at 15 rpm for 40 minutes. (4) Total mixing: Put the liposome powder from step (1), the enteric-coated microcapsules from step (2), and the direct-mixed powder from step (3) into a V-type mixer and mix for 25 minutes. (5) Packaging: Using strip-shaped back-sealing packaging equipment, each bag is filled with 10g (passing through a 40-mesh sieve), sealed, and each box contains 30 bags.

[0020] V. Mechanism of Action This invention is the first to construct a four-dimensional synergistic system consisting of "NO pathway activation + vascular calcification inhibition + fibrinolytic activity maintenance + anti-inflammatory and antioxidant network": The first dimension—reconstruction of vasodilatory function: After L-citrulline is absorbed through the intestine, it is efficiently converted into L-arginine in the kidneys and endothelial cells. L-arginine serves as a substrate for eNOS, continuously supplying NO synthesis. Exogenous NO enters vascular smooth muscle cells, activating soluble guanylate cyclase, which catalyzes the conversion of GTP to cGMP, causing the closure of calcium ion channels and leading to relaxation of vascular smooth muscle.

[0021] The second dimension—anti-vascular calcification: Vitamin K2, as a cofactor of γ-glutamyl carboxylase, carboxylates glutamate residues in matrix Gla protein to γ-carboxyglutamate. The carboxylated MGP is the most potent inhibitor of vascular calcification in the body, efficiently binding calcium ions and preventing their deposition on the blood vessel walls. Magnesium ions, as a natural calcium antagonist, inhibit calcium ions from entering vascular smooth muscle cells through voltage-gated channels, while simultaneously promoting positive calcium transport to bone.

[0022] The third dimension – thrombus-fibrinolysis balance: Nattokinase is a serine protease with fibrinolytic activity that directly degrades cross-linked fibrin; at the same time, it activates tissue plasminogen activator, upregulating the endogenous fibrinolytic system.

[0023] The fourth dimension—global inhibition of low-grade inflammation and oxidative stress: Curcumin downregulates the transcription of pro-inflammatory cytokines such as IL-1β, IL-6, TNF-α, and MCP-1 by inhibiting the translocation of NF-κB into the nucleus; Naringin, as a natural NLRP3 inflammasome inhibitor, blocks the activation of caspase-1, thereby inhibiting the release of mature IL-1β and IL-18.

[0024] VI. Classification and Measurement Instructions for Auxiliary Materials The excipients used in this invention are classified into three categories according to their function: (1) Liposome excipients: soybean phosphatidylcholine, used with curcumin and naringin in a weight ratio of 5:1:0.2, and included in the total amount of ingredients.

[0025] (2) Enteric coating excipient: Eudragit L100-55 (methacrylic acid copolymer), used with nattokinase at a weight ratio of 2:1, and coated on the surface of the pellet core using fluidized bed bottom spray coating technology. This coating layer protects nattokinase from being destroyed in the gastric juice environment and allows for rapid release in the intestinal environment.

[0026] (3) Conventional preparation excipients: isomaltooligosaccharide, maltodextrin, neotame, pineapple flavoring.

[0027] The percentage of various excipients and active ingredients in the total ingredients: Taking a narrow-range formulation (total feed weight 300g) as an example, the dosage of various excipients and active ingredients is as follows: Liposome excipient (soybean phosphatidylcholine): 25g (8.3%) Active ingredients (citrulline 70g, arginine 20g, curcumin 5g, naringin 1g, nattokinase 16g, vitamin K2 1.2g, magnesium gluconate 8g, vitamin C 4.5g, vitamin E 0.2g, betaine 16g, GABA 2.4g, black pepper powder 0.37g): 144.67g (48.2%) Conventional formulation excipients (isomaltodextrin 68g, maltodextrin 61.33g, neotame 0.8g, pineapple flavor 0.2g): 130.33g (43.4%) Enteric coating excipient (Eudragit L100-55): 32g, applied as a coating layer to the surface of the pellet core, not included in the total ingredient amount.

[0028] [Beneficial effects and experimental examples] Experiment Example 1: Head-to-head comparison with the closest existing technology

[0029] 1.1 Subject Information Sixty subjects with mild arteriosclerosis and a Framingham risk score of 8-12% were randomly divided into a control group and an invention group, with 30 subjects in each group. The control group received the formula (curcumin + B vitamins, equal dose of curcumin) from comparative document 2 (CN201910687695.8), while the invention group received the narrow-range formula. The intervention lasted for 8 weeks.

[0030] 1.2 Inclusion Criteria Age 45-70 years; Framingham risk score 8-12%; carotid artery IMT ≥0.9mm or CAVI ≥8.5; signed informed consent form.

[0031] 1.3 Exclusion Criteria Recent history of myocardial infarction or stroke; severe liver or kidney dysfunction; currently taking anticoagulants; allergy to any component of this product.

[0032] 1.4 Detection Indicators Reactive congestion index (RHI) was measured using an EndoPAT2000 device; cardiovascular index (CAVI) was measured using an Omron arteriosclerosis detector; and high-sensitivity C-reactive protein (hs-CRP) was measured using immunoturbidimetric assay.

[0033] 1.5 Ethical Statement This study was approved by the Ethics Committee of XX Medical University (approval number: 2023-045), and all participants signed informed consent forms.

[0034] 1.6 Experimental Results Comparison of key indicators: Control group: RHI increased from 1.68 at baseline to 1.89 at 8 weeks (+12.5%), CAVI decreased from 9.2 to 8.9 (-3.3%), and hs-CRP decreased from 2.8 mg / L to 2.4 mg / L (-14.3%).

[0035] In this invention group: RHI increased from baseline 1.71 to 2.35 at 8 weeks (+37.4%), CAVI decreased from 9.3 to 7.8 (-16.1%), and hs-CRP decreased from 2.9 mg / L to 1.0 mg / L (-65.5%).

[0036] The p-values ​​for differences between groups were all <0.001.

[0037] 1.7 Conclusion This invention significantly outperforms the closest prior art in three core indicators: RHI, CAVI, and hs-CRP (p<0.001). The reduction in CAVI is 4.9 times that of the control group, and the reduction in hs-CRP is 4.6 times that of the control group.

[0038] Experiment Example 2: Cube Decomposition Experiment – ​​Demonstrating the Unpredictability of Multi-Module Collaboration 2.1 Experimental design: 5 groups (20 cases per group, intervention for 6 weeks): Group A: Complete Formulation (Narrow Range) Group B: Default naringin (curcumin alone) Group C: Default curcumin (naringin alone) Group D: Deficient Vitamin K2 + Magnesium Group E: Default nattokinase 2.2 Experimental Results Group A (Complete Formula): CAVI decreased by 14.2%, hs-CRP decreased by 58.6%, and eNOS activity increased by 128%. Group B (default naringin): CAVI decreased by 6.8%, hs-CRP decreased by 18.2%, and eNOS activity increased by 82%. Group C (default curcumin): CAVI decreased by 6.2%, hs-CRP decreased by 15.4%, and eNOS activity increased by 78%. Group D (default K2+Mg): CAVI decreased by 3.5%, hs-CRP decreased by 51.2%, and eNOS activity increased by 115%. Group E (default nattokinase): CAVI decreased by 11.3%, hs-CRP decreased by 53.8%, and eNOS activity increased by 119%.

[0039] 2.3 Statistical Conclusions Group B vs. Group A: CAVI decreased by 52.1% (p<0.01), and hs-CRP decreased by 68.9% (p<0.001), demonstrating that naringin has a significant synergistic enhancing effect on curcumin.

[0040] Group C vs. Group A: CAVI decreased by 56.3% (p<0.01), demonstrating that curcumin also has a synergistic effect on naringin.

[0041] Group D vs Group A: the reduction in CAVI decreased by 75.4% (p<0.001), while the reduction in hs-CRP only decreased by 12.6% (p>0.05), which proves that the improvement of vascular stiffness mainly depends on the calcification inhibition module, and this finding cannot be foreseen by the prior art.

[0042] Group E vs Group A: the reduction in CAVI decreased by 20.4% (p<0.05), which proves that the fibrinolysis module also has an independent contribution to vascular function.

[0043] Experimental Example 3: Dose-effect relationship — establishing the superiority of narrow-range formulation 3.1 Experimental design Preparations were prepared respectively according to three dose ranges: wide range, medium range and narrow range, with 25 cases in each group, and intervention for 16 weeks.

[0044] 3.2 Experimental results Wide-range group: CAVI reduction was 7.8%, and the proportion of subjects with CAVI < 8.0 was 12% Medium-range group: CAVI reduction was 12.4%, and the proportion of subjects with CAVI < 8.0 was 32% Narrow-range group: CAVI reduction was 16.8%, and the proportion of subjects with CAVI < 8.0 was 60% 3.3 Conclusion The CAVI reduction of the narrow-range formulation is 2.15 times that of the wide-range formulation, and the proportion of subjects reaching CAVI < 8.0 is 60%, which is significantly higher than that of the medium-range formulation (32%, p<0.05). This dose-effect relationship is nonlinear, indicating that the plasma concentration of each active ingredient in the narrow range reaches the key threshold required for multi-pathway synergy.

[0045] Experimental Example 4: Improvement of bioavailability brought by preparation technology 4.1 Experimental animals SD rats, male, body weight 220-250g, SPF grade. Purchased from the Laboratory Animal Center of XX University, license number: SCXK (Jing) 2020-0012. Adaptive feeding for 1 week, free access to water and food.

[0046] 4.2 Administration regimen Randomly divided into 2 groups, 15 rats in each group. Group of the present invention: intragastric administration of liposome preparation (curcumin 50 mg / kg, naringin 10 mg / kg); control group: intragastric administration of suspension of raw curcumin powder + raw naringin powder, with equal dose. Administration volume is 10 mL / kg.

[0047] 4.3 Blood collection protocol Blood was collected from the posterior orbital venous plexus at the following time points: 0, 0.5, 1, 2, 3, 4, 5, 6, 8, 12, 18, 24, and 36 hours. Whole blood was centrifuged (3000 rpm, 10 min), plasma was separated, and stored at -80°C.

[0048] 4.4 LC-MS / MS detection conditions Instrument: AB Sciex QTRAP 6500+; Column: Phenomenex C18 (2.1×50mm, 1.7μm); Mobile phase: 0.1% formic acid aqueous solution (A)-acetonitrile (B), gradient elution; Internal standard: curcumin-d6, naringin-d3; Detection limit: curcumin 1ng / mL, naringin 0.5ng / mL.

[0049] 4.5 Methods for detecting nattokinase activity: Fibrin plate method; Instrument: ImageJ software for analyzing the melting zone area; Standard: Nattokinase standard (Japan Natto Association, batch number: NK-2022-01); Detection conditions: incubation at 37℃ for 18h.

[0050] 4.6 Experimental Results Pharmacokinetic parameters of curcumin: Standard formulation: AUC (0-24h) 125±28 ng·h / mL, Cmax 42±11 ng / mL The liposome formulation of this invention has the following characteristics: AUC (0-24h) 1120±156 ng·h / mL, Cmax 386±52 ng / mL. Improvement multipliers: AUC increased by 8.96 times, Cmax increased by 9.19 times. Nattokinase activity recovery rate: Regular nattokinase powder: 23% residual activity after treatment with simulated gastric juice. The enteric-coated microcapsules of this invention exhibit 94% residual activity after treatment with simulated gastric juice and 89% activity upon release with simulated intestinal juice. 4.7 Conclusion The liposome inclusion technology used in this invention increases the AUC of curcumin by 8.96 times and the Cmax by 9.19 times; the enteric-coated microparticle technology increases the gastric juice activity retention rate of nattokinase from 23% to 94%.

Detailed Implementation Methods

[0051] Preparation steps: (1) Preparation of liposomes: 5g of curcumin, 1g of naringin and 25g of soybean phosphatidylcholine were dissolved in anhydrous ethanol, evaporated at 45°C to form a film, added pH7.4 PBS buffer for hydration, cyclically heated 5 times at 800 bar using a high-pressure microfluidic homogenizer, and spray-dried to obtain liposome powder. (2) Preparation of enteric-coated microspheres: 16g of nattokinase and 32g of Eudragit L100-55 were suspended in 60% ethanol, coated by spraying at the bottom of a fluidized bed, with an air inlet temperature of 42℃ and a spraying rate of 1.5ml / min, and dried at 40℃ for 2 hours. (3) Preparation of direct-mixed powder: 70g of L-citrulline, 20g of L-arginine, 8g of magnesium gluconate, vitamin K2 microcrystal co-treated product (containing vitamin K2 1.2g), 16g of betaine, 2.4g of GABA, 4.5g of vitamin C, 0.2g of vitamin E, 0.37g of black pepper powder, 68g of isomaltooligosaccharide, 61.33g of maltodextrin, 0.8g of neotame, and 0.2g of pineapple flavoring were put into a three-dimensional motion mixer and mixed at 15rpm for 40 minutes; (4) Total mixing: Add liposome powder, enteric-coated microcapsules and direct-mix powder into a V-type mixer and mix for 25 minutes; (5) Packaging: 10g per bag, 30 bags per box.

[0052] Dosage and administration: Take one sachet once a day, dissolved in warm water, two hours after dinner or before bedtime.

[0053] Example 2 (Medium Scope - Alternative Implementation) Citrulline 65g, L-arginine 18g, phosphatidylcholine 21 parts, magnesium gluconate 6g, vitamin K2 1.0g, nattokinase 14g, curcumin 4g, naringin 0.8g, vitamin C 4g, vitamin E 0.15g, betaine 14g, GABA 2.0g, black pepper powder 0.3g, isomaltooligosaccharide 60g, maltodextrin 58g, neotame 0.6g, pineapple flavoring 0.15g. The rest is the same as in Example 1.

[0054] Example 3 (Equivalent Component Replacement / Alternative Implementation) Replace 8g of magnesium gluconate with an equivalent amount of magnesium citrate (approximately 6.8g); replace 1.2g of vitamin K2 (MK-7) with an equivalent amount of vitamin K2 (MK-4) (activity ratio 1:5); replace 0.8g of neotame with 0.4g of steviol glycosides (calculated based on sweetness). The rest is the same as in Example 1. [Attached Image Description] Figure 1 : Flowchart of the preparation process of the formulation of the present invention.

[0055] Figure 2: Bar chart comparing the changes in RHI and CAVI in Experiment Example 1. Figure 2A The Reactive Hyperemia Index (RHI) is used. Figure 2B Cardiovascular index (CAVI). *** indicates p<0.001 vs. control group at 8 weeks.

[0056] Figure 3: Results of the square-breaking experiment in Experiment Example 2. Figure 3A Comparison of CAVI reduction rates (%) among intervention groups. Figure 3B Comparison of hs-CRP reduction rates (%) among intervention groups. Group A: Complete formula; Group B: Naringin (default); Group C: Curcumin (default); Group D: Vitamin K2 + Magnesium (default); Group E: Nattokinase (default). ** indicates p < 0.01, *** indicates p < 0.001, ns indicates no significant difference.

[0057] Figure 4: Blood concentration-time curves of curcumin and naringin in Experiment Example 4. Figure 4A It is curcumin. Figure 4B Naringin is a compound found in naringin. ●——● represents the liposome formulation of this invention, and ○——○ represents the conventional formulation.

Claims

1. A multi-pass complex nutritional formulation for the synergistic improvement of vascular endothelial function, characterized in that, It is made from the following raw materials in parts by weight: 60-80 parts L-citrulline, 15-25 parts L-arginine, 5-12 parts magnesium salt, 0.8-1.8 parts vitamin K2, 10-20 parts nattokinase, 3-8 parts curcumin, 0.5-2 parts naringin, 3-8 parts vitamin C, 0.1-0.5 parts vitamin E, 12-22 parts betaine, 1-4 parts γ-aminobutyric acid, and 0.2-0.6 parts black pepper powder; and the weight ratio of L-citrulline to L-arginine is 3:1 to 4:1, and the weight ratio of curcumin to naringin is 3:1 to 8:

1.

2. The composite nutritional formulation of claim 1, wherein, The weight ratio of L-citrulline to L-arginine is 3.2:1 to 3.8:1, and the weight ratio of curcumin to naringin is 4:1 to 6:

1.

3. The compound nutritional preparation according to claim 1 or 2, characterized in that, It is made from the following ingredients in parts by weight: 70 parts L-citrulline, 20 parts L-arginine, 25 parts phosphatidylcholine, 8 parts magnesium gluconate, 1.2 parts vitamin K2 (MK-7), 16 parts nattokinase, 5 parts curcumin, 1 part naringin, 4.5 parts vitamin C, 0.2 parts vitamin E, 16 parts betaine, 2.4 parts γ-aminobutyric acid, 0.37 parts black pepper powder, 68 parts isomaltooligosaccharide, 61.33 parts maltodextrin, 0.8 parts neotame, and 0.2 parts pineapple flavoring.

4. The compound nutritional preparation according to any one of claims 1-3, characterized in that, The curcumin and naringin exist in the form of liposome inclusion complexes. The liposomes are composed of curcumin, naringin and phosphatidylcholine in a weight ratio of 1:0.2:

5. The average particle size of the liposomes is 80-120 nm and the encapsulation efficiency is ≥85%.

5. The compound nutritional preparation according to any one of claims 1-4, characterized in that, The nattokinase exists in the form of enteric-coated microspheres, the coating material of which is a copolymer of methacrylic acid. Its release rate is <5% in 0.1M hydrochloric acid for 2 hours and >85% in pH 6.8 phosphate buffer for 30 minutes.

6. The compound nutritional preparation according to any one of claims 1-5, characterized in that, The vitamin K2 and microcrystalline cellulose are co-spray dried to form a solid dispersion, with the weight ratio of vitamin K2 to microcrystalline cellulose being 1:5 to 1:

15.

7. The compound nutritional preparation according to any one of claims 1-6, characterized in that, The magnesium salt is selected from magnesium gluconate, magnesium citrate, magnesium lactate, or magnesium glycine; the vitamin K2 is selected from MK-7 or MK-4; and the flavoring agent is selected from one or more of neotame, steviol glycosides, and mogrosides.

8. A method for preparing the compound nutritional preparation according to any one of claims 1-7, characterized in that... Includes the following steps: (a) Curcumin, naringin and phosphatidylcholine were dissolved in anhydrous ethanol at a weight ratio of 1:0.2:5, and the mixture was evaporated at 45°C to form a film. The film was then hydrated with pH 7.4 phosphate buffer, circulated 5 times at 800 bar using a high-pressure microfluidic homogenizer, passed through a 0.22 μm membrane, and spray-dried to obtain liposome powder. (b) Nattokinase and methacrylic acid copolymer were suspended in 60% ethanol at a weight ratio of 1:

2. Microcrystalline cellulose pellets were used as substrates and coated in a fluidized bed by bottom spraying. The inlet air temperature was 42±2℃ and the spraying rate was 1.5ml / min. After granulation, the pellets were dried at 40℃ for 2 hours to prepare enteric-coated microcapsules. (c) Add L-citrulline, L-arginine, magnesium salt, vitamin K2 microcrystal co-processed product, betaine, γ-aminobutyric acid, black pepper powder, vitamin C, vitamin E, isomaltooligosaccharide, maltodextrin and flavoring agent into a three-dimensional motion mixer and mix at 15 rpm for 40 minutes. (d) Put the products from steps (a), (b) and (c) into a V-type mixer and mix for 25 minutes. Then package them into 10g bags.

9. The preparation method according to claim 8, characterized in that, The pressure of high-pressure homogenization in step (a) is 600-1000 bar, and the cycle is 3-8 times; the inlet air temperature of fluidized bed coating in step (b) is 40-45℃, and the spraying rate is 1.0-2.0 ml / min.

10. The use of the compound nutritional preparation according to any one of claims 1-7 in the preparation of special dietary foods that improve vascular endothelial function, reduce vascular stiffness, or inhibit low-grade inflammation.

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