High-activity fermented astragalus mongholicus and red date compound for improving myocardial function as well as preparation method and application thereof

Abstract

The invention relates to the technical field of food biology, in particular to a high-activity fermented astragalus mongholicus and red date compound for improving myocardial function, which is prepared by taking Xinjiang Qiang red dates and astragalus mongholicus as main raw materials and matching cellulase, pectinase, beta-glucanase and hydroxypropyl-beta-cyclodextrin. In the preparation process, gradient freezing-microwave pretreatment and ternary composite enzymolysis synergistic wall breaking are adopted, quaternary strains of bifidobacterium longum, bacillus subtilis, saccharomyces cerevisiae and lactobacillus plantarum are compounded, adenosine concentration response type staged solid-state fermentation is performed, and microcapsule embedding stabilizing treatment is performed to obtain the product. The content of cAMP in the product is greater than or equal to 13.0 mu g / g, and the content of calycosin-7-glucoside in astragalus membranaceus is greater than or equal to 1.9 mg / g. The compound can be used for preparing myocardial ischemia pretreatment health-care food and a pharmaceutical composition for adjuvant therapy in the myocardial infarction recovery period, and the dosage forms comprise chewable tablets, oral liquid, capsules and the like.

Description

Technical Field

[0001] This invention relates to the field of food biotechnology, specifically to a highly active fermented astragalus and jujube complex for improving myocardial function, its preparation method, and its application. Background Technology

[0002] Myocardial ischemic disease is a major chronic disease that threatens human health worldwide. Its pathogenesis is closely related to myocardial cell hypoxia, energy metabolism disorders, apoptosis, and oxidative stress damage. Astragalus and jujube, as traditional medicinal and edible ingredients, are often used in traditional Chinese medicine to invigorate qi and blood circulation, nourish the heart and calm the mind. Modern pharmacological studies have confirmed that saponins in astragalus (such as verrucoside), flavonoids in jujube (such as 6,8-diglucosylnaringenin), and cyclic adenosine monophosphate (cAMP) shared by both can exert cardioprotective effects by regulating myocardial cell signaling pathways, inhibiting oxidative stress, and reducing myocardial cell apoptosis.

[0003] However, the active ingredients in natural astragalus and jujube suffer from low dissolution rates and poor bioavailability. The cAMP content in untreated raw materials is only 0.52±0.03μg / g, the total saponin dissolution rate of astragalus is less than 50%, and the flavonoids in jujube are difficult to release due to cell wall encapsulation. Microbial fermentation is the mainstream technology for enhancing the activity of medicinal and edible raw materials, but existing related technologies mostly use single-strain (such as Lactobacillus plantarum) or binary-strain fermentation, with cAMP enhancement generally less than 12-fold and verbascoside glucoside content less than 0.4mg / g, far from meeting the clinical-grade activity requirements; furthermore, relying on mechanical pulverization or single enzymatic hydrolysis results in a cell wall destruction rate of ≤75%, insufficient release of precursor substances (such as adenosine and saponin precursors), and limited transformation efficiency of the strain.

[0004] Therefore, developing an Astragalus and Jujube fermentation technology that can achieve high activity enrichment, high cell wall disruption efficiency, high stability, and high reproducibility, while clarifying its application scenarios in cardioprotection, is of great significance for promoting the industrialization of functional foods and adjuvant therapeutic drugs. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a highly active fermented astragalus and jujube complex for improving myocardial function, its preparation method, and its application, solving the problems of low enrichment efficiency of active ingredients, incomplete cell wall disruption, and poor product storage stability in existing technologies.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution: a highly active fermented astragalus and jujube complex for improving myocardial function, comprising the following raw materials in parts by weight: 60-80 parts of Xinjiang Ruoqiang jujube, 20-40 parts of astragalus, 0.1-1.0 parts of cellulase, 0.1-1.0 parts of pectinase, 0.1-1.0 parts of β-glucanase, and 0.1-1.0 parts of hydroxypropyl-β-cyclodextrin.

[0007] Preferably, a method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function includes the following steps: a) Gradual freezing-thawing treatment of raw materials Select Xinjiang Ruoqiang red dates with a sugar content ≥65 Brix, remove the pits, and dry them in hot air at 30-40℃ until the moisture content is ≤12%. Cut them into 1-2cm pieces. 3 Small pieces; take Astragalus membranaceus with a basic saponin content ≥2.0mg / g, slice it to a thickness of 2-3mm, and pre-treat it with microwave at 2450MHz and 300W for 1-2min; place the above-mentioned jujube pieces and Astragalus membranaceus slices separately at -40℃ for 1h, then transfer them to ≤-85℃ for rapid freezing for 2-3 hours, take them out and thaw them at 20-25℃ with circulating air for 1-1.5 hours, repeat this gradient freezing-thawing process twice; after treatment, pulverize them separately, jujube powder through an 80-mesh sieve, and Astragalus membranaceus powder through a 60-mesh sieve. At this time, the cell destruction rate of jujube is ≥98.5%, and the cell destruction rate of Astragalus membranaceus is ≥96.2%; b) Preparation of composite matrix The frozen jujube powder obtained in step a) is mixed with astragalus powder at a mass ratio of (6-8):(2-4) to obtain a composite matrix; c) Pretreatment with ternary compound enzymatic hydrolysis Cellulase (50 U / g), pectinase (35 U / g), and β-glucanase (15 U / g) were added to the composite matrix, and hydrolysis was carried out at 48-52℃ and pH 4.8-5.2 for 1.5-2.5 hours. The enzymatic hydrolysis, combined with the gradient freezing treatment in step a), resulted in a matrix cell wall destruction rate of ≥99.1%. d) Substrate sterilization Add water to the enzymatically hydrolyzed matrix to a moisture content of 56%-59%, autoclave at 121℃ for 18-20 minutes, and cool to room temperature; e) Activation and compounding of tetrad strains Bifidobacterium longum was inoculated into MRS liquid medium and cultured anaerobically at 37°C until OD200. 600 =0.6-0.8, centrifuged and resuspended in sterile physiological saline to 1×10⁻⁶. 8 CFU / mL, β-glucosidase activity ≥112.3 U / g; Bacillus subtilis inoculated into LB liquid medium and cultured in a shaker at 37°C until OD... 600 =0.8-1.0, bacterial concentration 1×108 CFU / mL, protease activity ≥98.7 U / g; Saccharomyces cerevisiae inoculated into YPD liquid medium and cultured at 37℃ until OD 600 =0.6-0.8, bacterial concentration 1×10 8 CFU / mL, sucrase activity ≥135.2 U / g; *Lactobacillus plantarum* inoculated into MRS liquid medium and incubated statically at 37°C until OD500. 600 =0.7-0.9, bacterial concentration 1×10 8 CFU / mL, cAMP synthase activity ≥90.6U / g; four bacterial solutions were mixed in a volume ratio of (1.5-2.5):(1.5-2.5):(0.8-1.2):(0.8-1.2) and sprayed onto the substrate obtained in step d) at a total inoculum of 3%-4% (v / w); f) Adenosine concentration-responsive staged solid-state fermentation Under the condition of real-time monitoring of adenosine concentration in fermentation broth, the following parameters were controlled in stages: 0-10h: Temperature 38-39℃, pH 6.7-6.9, DO 6-10%. Maintain this stage when adenosine concentration ≤0.5mg / L to promote the proliferation of Bacillus subtilis and Lactobacillus plantarum. 10-36h: Temperature 36-37℃, pH 6.4-6.6, DO 0.5%-1%. When the adenosine concentration is ≥0.8mg / L, this stage is entered, which is the main fermentation stage, and the cAMP synthesis rate is ≥0.37μg / (g・h). 36-60h: Temperature 34-35℃, pH 6.2-6.4, DO < 0.3%, fermentation ends when the content of verbascoside isoflavone glucoside is ≥ 1.8 mg / g; g) Post-processing for product stabilization After fermentation, the mixture was freeze-dried at -50℃ until the moisture content was ≤5%, pulverized and passed through a 100-mesh sieve, and then microencapsulated with 10% (w / w) hydroxypropyl-β-cyclodextrin (inlet air temperature 180℃, outlet air temperature 80℃, spray rate 5mL / min) to obtain the fermented astragalus and jujube complex.

[0008] Preferably, in step a), the microwave pretreatment parameters are 2450MHz, 300W, and 1.5min. After gradient freezing and thawing, the cell destruction rate of jujube reaches 98.7±1.2%, and the cell destruction rate of astragalus reaches 96.5±1.5%. In step c), the hydrolysis conditions were 50℃ and pH 5.0 for 2 hours. After hydrolysis by the ternary complex enzyme, the cell wall destruction rate reached 99.3±0.8%.

[0009] Preferably, in step e), the inoculation volume ratio of the four-component bacterial solution is 2:2:1:1, the total inoculation amount is 3.5% (v / w), and the total number of viable bacteria in the substrate after inoculation is ≥3×107 CFU / g; the Lactobacillus plantarum is a Gram-positive facultative anaerobic bacterium with acid-resistant characteristics.

[0010] Preferably, the termination condition for responsive fermentation in step f) is: cAMP content ≥ 13.0 μg / g and 6,8-diglucosyl-naringenin content ≥ 6.7 mg / g, at which point the total fermentation cycle is 48 hours.

[0011] Preferably, the hydroxypropyl-β-cyclodextrin encapsulated in the microcapsules in step g) has a purity of ≥99%, and the encapsulated complex has a solubility of 18.5 mg / mL at 25 °C.

[0012] This invention provides a highly active fermented astragalus and jujube complex for improving myocardial function, its preparation method, and its application. It possesses the following beneficial effects: 1. The fermented Astragalus and Jujube complex of this invention contains ≥13.0 μg / g (dry weight) cAMP, which is ≥25 times higher than that of unfermented raw materials and 138.1% higher than the existing best technology. The content of Astragalus isoflavone glucoside is ≥1.9 mg / g, which is 9.5 times the value specified in the Chinese Pharmacopoeia (0.2 mg / g). The content of 6,8-digluxyl-2(S) / 2(R)-naringenin in jujube is ≥6.7 mg / g, which is 13.4 times higher than that of unfermented raw materials.

[0013] 2. This invention achieves a matrix cell wall destruction rate of ≥99.1% through the synergistic effect of gradient freezing-microwave disruption and ternary compound enzymatic hydrolysis, which is 32.4% higher than the 75% of existing single enzymatic hydrolysis; the total saponin dissolution rate of Astragalus membranaceus reaches 85.2%, providing sufficient precursor substrates for subsequent strain transformation.

[0014] 3. This invention uses hydroxypropyl-β-cyclodextrin microcapsules for encapsulation. After 3 months of storage at room temperature and away from light, the cAMP degradation rate is ≤2.3%, which is more than 78% lower than the 14.8% of unencapsulated products and the 13.7% of existing products, thus solving the industry problem of unstable cAMP storage. Attached Figure Description

[0015] Figure 1 This image shows a comparison of the cell disruption effects of gradient freezing-microwave pretreatment and single freezing according to the present invention. The left side represents single freezing with a cell integrity rate of 12%, while the right side represents gradient freezing-microwave pretreatment with a cell integrity rate of 1.3%. Figure 2 This is a line graph showing the relationship between the number of freeze-thaw cycles and the cAMP boost factor in this invention. Figure 3 This is a bar chart comparing the cAMP content and enhancement factor of the present invention with those of existing technologies. Figure 3 A represents a comparison of increased cAMP levels. Figure 3 B represents the comparison of the multiplier. Detailed Implementation

[0016] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Example 1

[0017] This invention provides a highly active fermented astragalus and jujube complex for improving myocardial function, comprising the following raw materials in parts by weight: 80 parts of Xinjiang Ruoqiang jujube, 40 parts of astragalus, 1.0 part of cellulase, 1.0 part of pectinase, 1.0 part of β-glucanase, and 1.0 part of hydroxypropyl-β-cyclodextrin.

[0018] A method for preparing a highly active fermented astragalus and jujube complex that improves myocardial function includes the following steps: a) Gradual freezing-thawing treatment of raw materials Select Xinjiang Ruoqiang red dates with a sugar content ≥65 Brix, remove the pits, dry them in hot air at 40℃ until the moisture content is ≤12%, and cut them into 2cm pieces. 3 Small pieces; take Astragalus membranaceus with a basic saponin content ≥2.0mg / g, slice it to a thickness of 3mm, and pre-treat it with microwave at 2450MHz and 300W for 2min. The microwave pre-treatment parameters are 2450MHz, 300W, and 1.5min. Place the above-mentioned jujube pieces and Astragalus membranaceus slices at -40℃ for 1h, and then transfer them to ≤-85℃ for rapid freezing for 3 hours. After taking them out, thaw them at 25℃ with circulating air for 1.5 hours. Repeat this gradient freezing-thawing process twice. After treatment, grind them separately. Jujube powder is passed through an 80-mesh sieve, and Astragalus membranaceus powder is passed through a 60-mesh sieve. At this time, the cell destruction rate of jujube is ≥98.5%, and the cell destruction rate of Astragalus membranaceus is ≥96.2%. b) Preparation of composite matrix The frozen jujube powder obtained in step a) is mixed with astragalus powder at a mass ratio of 8:4 to obtain a composite matrix; c) Pretreatment with ternary compound enzymatic hydrolysis Cellulase (50 U / g), pectinase (35 U / g), and β-glucanase (15 U / g) were added to the composite matrix, and hydrolysis was carried out at 52℃ and pH 5.2 for 2.5 hours. The enzymatic hydrolysis and the gradient freezing treatment in step a) synergistically resulted in a matrix cell wall destruction rate of ≥99.1%. d) Substrate sterilization Add water to the enzymatically hydrolyzed matrix to a moisture content of 59%, autoclave at 121°C for 20 minutes, and cool to room temperature; e) Activation and compounding of tetrad strains Bifidobacterium longum was inoculated into MRS liquid medium and cultured anaerobically at 37°C until OD200. 600 =0.8, centrifuged and resuspended in sterile saline to 1×10⁻⁸. 8 CFU / mL, β-glucosidase activity ≥112.3 U / g; Bacillus subtilis inoculated into LB liquid medium and cultured in a shaker at 37°C until OD... 600 =1.0, bacterial concentration 1×10 8 CFU / mL, protease activity ≥98.7 U / g; Saccharomyces cerevisiae inoculated into YPD liquid medium and cultured at 37℃ until OD 600 =0.6-0.8, bacterial concentration 1×10 8 CFU / mL, sucrase activity ≥135.2 U / g; *Lactobacillus plantarum* inoculated into MRS liquid medium and incubated statically at 37°C until OD500. 600 =0.7-0.9, bacterial concentration 1×10 8 CFU / mL, cAMP synthase activity ≥90.6 U / g; four bacterial solutions were mixed in a volume ratio of 2.5:2.5:1.2:1.2 and sprayed onto the substrate obtained in step d) at a total inoculum of 4% (v / w); f) Adenosine concentration-responsive staged solid-state fermentation Under the condition of real-time monitoring of adenosine concentration in fermentation broth, the following parameters were controlled in stages: 0-10h: Temperature 39℃, pH 6.9, DO 10%. Maintain this stage when adenosine concentration ≤ 0.5 mg / L to promote the proliferation of Bacillus subtilis and Lactobacillus plantarum. 10-36h: Temperature 37℃, pH -6.6, DO 1%. When the adenosine concentration is ≥0.8mg / L, this stage is entered, which is the main fermentation stage, and the cAMP synthesis rate is ≥0.37μg / (g・h). 36-60h: Temperature 35℃, pH 6.4, DO < 0.3%, fermentation ends when the content of verbascoside isoflavone glucoside is ≥ 1.8 mg / g; g) Post-processing for product stabilization After fermentation, the mixture was freeze-dried at -50℃ until the moisture content was ≤5%, pulverized and passed through a 100-mesh sieve, and then microencapsulated with 10% (w / w) hydroxypropyl-β-cyclodextrin (inlet air temperature 180℃, outlet air temperature 80℃, spray rate 5mL / min) to obtain the fermented astragalus and jujube complex. Example 2

[0019] This invention provides a highly active fermented astragalus and jujube complex for improving myocardial function, comprising the following raw materials in parts by weight: 60 parts of Xinjiang Ruoqiang jujube, 20 parts of astragalus, 0.1 parts of cellulase, 0.1 parts of pectinase, 0.1 parts of β-glucanase, and 0.1 parts of hydroxypropyl-β-cyclodextrin.

[0020] A method for preparing a highly active fermented astragalus and jujube complex that improves myocardial function includes the following steps: a) Gradual freezing-thawing treatment of raw materials Select Xinjiang Ruoqiang red dates with a sugar content ≥65 Brix, remove the pits, and dry them in hot air at 30℃ until the moisture content is ≤12%. Cut them into 1cm pieces. 3 Small pieces; take Astragalus membranaceus with a basic saponin content ≥2.0mg / g, slice it to a thickness of 2mm, and pre-treat it with microwave at 2450MHz and 300W for 1min. The microwave pre-treatment parameters are 2450MHz, 300W, and 1.5min. Place the above-mentioned jujube pieces and Astragalus membranaceus slices at -40℃ for 1h, and then transfer them to ≤-85℃ for rapid freezing for 2 hours. After taking them out, thaw them at 20℃ with circulating air for 1 hour. Repeat this gradient freezing-thawing process twice. After treatment, grind them separately. Jujube powder is passed through an 80-mesh sieve, and Astragalus membranaceus powder is passed through a 60-mesh sieve. At this time, the cell destruction rate of jujube is ≥98.5%, and the cell destruction rate of Astragalus membranaceus is ≥96.2%. b) Preparation of composite matrix The frozen jujube powder obtained in step a) was mixed with astragalus powder at a mass ratio of 6:2 to obtain a composite matrix; c) Pretreatment with ternary compound enzymatic hydrolysis Cellulase (50 U / g), pectinase (35 U / g), and β-glucanase (15 U / g) were added to the composite matrix, and hydrolysis was carried out at 48 °C and pH 4.8 for 1.5 hours. The enzymatic hydrolysis and the gradient freezing treatment in step a) synergistically resulted in a matrix cell wall destruction rate of ≥99.1%. d) Substrate sterilization Add water to the enzymatically hydrolyzed matrix to a moisture content of 56%, autoclave at 121°C for 18 minutes, and cool to room temperature; e) Activation and compounding of tetrad strains Bifidobacterium longum was inoculated into MRS liquid medium and cultured anaerobically at 37°C until OD200. 600 =0.6, centrifuged and resuspended in sterile saline to 1×10⁶. 8 CFU / mL, β-glucosidase activity ≥112.3 U / g; Bacillus subtilis inoculated into LB liquid medium and cultured in a shaker at 37°C until OD... 600 =0.8, bacterial concentration 1×10 8 CFU / mL, protease activity ≥98.7 U / g; Saccharomyces cerevisiae inoculated into YPD liquid medium and cultured at 37℃ until OD 600=0.6, bacterial concentration 1×10 8 CFU / mL, sucrase activity ≥135.2 U / g; *Lactobacillus plantarum* inoculated into MRS liquid medium and incubated statically at 37°C until OD500. 600 =0.7, bacterial concentration 1×10 8 CFU / mL, cAMP synthase activity ≥90.6 U / g; four bacterial solutions were mixed in a volume ratio of 1.5:1.5:0.8:0.8 and sprayed onto the substrate obtained in step d) at a total inoculum of 3% (v / w); f) Adenosine concentration-responsive staged solid-state fermentation Under the condition of real-time monitoring of adenosine concentration in fermentation broth, the following parameters were controlled in stages: 0-10h: Temperature 38℃, pH 6.7, DO 6%. Maintain this stage when adenosine concentration ≤ 0.5 mg / L to promote the proliferation of Bacillus subtilis and Lactobacillus plantarum. 10-36h: Temperature 36℃, pH 6.4, DO 0.5%. When the adenosine concentration is ≥0.8mg / L, this stage is entered, which is the main fermentation stage, and the cAMP synthesis rate is ≥0.37μg / (g・h). 36-60h: Temperature 34℃, pH 6.2, DO < 0.3%, fermentation ends when the content of verbascoside isoflavone glucoside is ≥ 1.8 mg / g; g) Post-processing for product stabilization After fermentation, the mixture was freeze-dried at -50℃ until the moisture content was ≤5%, pulverized and passed through a 100-mesh sieve, and then microencapsulated with 10% (w / w) hydroxypropyl-β-cyclodextrin (inlet air temperature 180℃, outlet air temperature 80℃, spray rate 5mL / min) to obtain the fermented astragalus and jujube complex.

[0021] Experiment 1: like Figure 1-2 As shown, this experiment provides a gradient freezing-thawing parameter optimization experiment to determine the optimal combination of freezing and thawing parameters: 1. Experimental Design Five treatment groups were set up, with 100g each of jujube pieces and astragalus slices in each group. Cell damage rate and adenosine content were measured after treatment. The gradient freeze-thaw treatment parameters for each group are shown in the table below: Table 1

[0022] 2. Operating steps (taking Group 4 as an example) Flash freezing: Spread the jujube pieces and astragalus slices evenly on a sterile tray and freeze in an ultra-low temperature freezer at -85℃ for 3 hours; Thawing at room temperature: After removing from the freezer, thaw in a constant temperature incubator at 25°C for 1.5 hours, turning the freezer over every 30 minutes during this period; Repeated processing: Repeat the freeze-thaw process once, then pulverize and test.

[0023] 3. Results Analysis The comparison of cell destruction rate and precursor release amount after freezing and thawing in each group is shown in the table below: Table 2

[0024] As shown in the table above, the optimal parameters are -85℃ freezing for 3 hours and thawing at 25℃ for 1.5 hours, repeated twice. At this time, the cell destruction rate and precursor release amount reach the optimal level, and further increasing the number of times is not cost-effective.

[0025] Experiment 2: This experiment provides verification of the synergistic effect of physical cell disruption and enzymatic hydrolysis, used to verify the synergistic effect of freezing treatment and enzymatic hydrolysis: 1. Experimental Design Group A: Enzymatic hydrolysis only (50℃, pH 5.0, hydrolysis for 2 hours); Group B: Rapid freeze-thaw (optimal parameters from Example 1) + no enzymatic hydrolysis; Group C: Rapid freezing and thawing + compound enzymatic hydrolysis (same as Group A).

[0026] 2. Results Analysis The cell wall disruption rate and cAMP content of each group are shown in the table below: Table 3

[0027] As shown in the table above, gradient freezing-thawing and ternary enzymatic hydrolysis have a significant synergistic effect, and the combined treatment maximizes both cell wall disruption rate and cAMP content.

[0028] Experiment 3: like Figure 3 As shown, this experiment compares the performance of this method with existing technologies to verify the differences: 1. Experimental Design This method group follows the process of group C in Example 2.

[0029] Control group 1: The process was carried out according to the process disclosed in Chinese Invention Patent Publication No. (CN118216611A), specifically: Clostridium butyricum-Lactobacillus plantarum-Saccharomyces boulardii = 1:1:1, fermentation for 72 hours.

[0030] Control group 2: Following the publicly disclosed process, the bacterial strain was adjusted to lactic acid bacteria-Lactobacillus plantarum-Slow-growing rhizobium = 1:1:1, and fermented for 72 hours.

[0031] 2. Results Analysis The cAMP content, active ingredient dissolution, and production efficiency of each group are shown in the table below: Table 4

[0032] As shown in the table above, the present invention is superior to the prior art in terms of cAMP content, dissolution of active ingredients, production efficiency, and product stability.

[0033] Experiment 4: This experiment verifies the effects of quaternary real bacterial strains combined with ternary bacterial strains: Experimental Design: Three strain combinations were set up with a total inoculum of 3.5%, and the characteristic components were detected after fermentation for 45 hours according to the preparation method of the present invention.

[0034] Group A (ternary): Bifidobacterium longum: Bacillus subtilis: Saccharomyces cerevisiae = 2:2:1; Group B (quaternary, this invention): Bifidobacterium longum: Bacillus subtilis: Saccharomyces cerevisiae: Lactobacillus plantarum = 2:2:1:1; Group C (according to Chinese Invention Patent Publication No. CN118216611A): Single Lactobacillus plantarum.

[0035] Results analysis: The results of the comparison between the effects of quaternary real bacterial strains and ternary bacterial strains are shown in the table below: Table 5

[0036] As shown in the table above, the quaternary functional strain compound system is significantly superior to the ternary strain combination and the existing single strain fermentation in all activity indicators, proving the necessity and superiority of strain synergy.

[0037] Experiment 5: This experiment verifies the stability of the microcapsule-encapsulated complex: Experimental Design: The complexes prepared by the method of this invention (Group 1, real bacterial strain + embedding), the unembedded complex (Group 2, real bacterial strain), and the product of Chinese Invention Patent Publication No. CN118216611A (Group 3, bacterial strain unknown) were stored at room temperature in the dark for 3 months, and the cAMP content was measured monthly. The results are shown in the table below: Table 6

[0038] As shown in the table above, the cAMP degradation rate after 3 months of encapsulation using the method of the present invention is only 2.3%, while the degradation rate of the unencapsulated group is 14.8%, and the degradation rate of existing products is 13.7%. Encapsulation treatment significantly improves storage stability; and because the strains used are traceable, the experimental results can be reproduced by third parties.

[0039] Experiment Six: This experiment verifies the cardioprotective effect of the complex prepared in this invention combined with tanshinone extract: Forty SPF-grade male ICR mice were randomly divided into four groups: sham-operated group, model group, the complex group of this invention (150 mg / kg, prepared with real bacterial strain), and the invention + Tanshinone extract group (150 mg / kg complex + 50 mg / kg Tanshinone extract). After coronary artery ligation to establish the model, the mice were administered the complex via gavage for 14 days, and the cardiomyocyte apoptosis rate was measured. The results are shown in the table below: Table 7

[0040] As shown in the table above, the complex of the present invention has a significant cardioprotective effect, and the effect is even better when combined with tanshinone extract.

[0041] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A highly active fermented astragalus and jujube complex for improving myocardial function, characterized in that: The ingredients include the following parts by weight: 60-80 parts of Xinjiang Ruoqiang red dates, 20-40 parts of Astragalus membranaceus, 0.1-1.0 parts of cellulase, 0.1-1.0 parts of pectinase, 0.1-1.0 parts of β-glucanase, and 0.1-1.0 parts of hydroxypropyl-β-cyclodextrin.

2. The method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function according to claim 1, characterized in that: Includes the following steps: a) Gradual freezing-thawing treatment of raw materials Select Xinjiang Ruoqiang red dates with a sugar content ≥65 Brix, remove the pits, and dry them in hot air at 30-40℃ until the moisture content is ≤12%. Cut them into 1-2cm pieces. 3 Small pieces; take Astragalus membranaceus with a basic saponin content ≥2.0mg / g, slice it to a thickness of 2-3mm, and pre-treat it with microwave at 2450MHz and 300W for 1-2min; place the above-mentioned jujube pieces and Astragalus membranaceus slices separately at -40℃ for 1h, then transfer them to ≤-85℃ for rapid freezing for 2-3 hours, take them out and thaw them at 20-25℃ with circulating air for 1-1.5 hours, repeat this gradient freezing-thawing process twice; after treatment, pulverize them separately, jujube powder through an 80-mesh sieve, and Astragalus membranaceus powder through a 60-mesh sieve. At this time, the cell destruction rate of jujube is ≥98.5%, and the cell destruction rate of Astragalus membranaceus is ≥96.2%; b) Preparation of composite matrix The frozen jujube powder obtained in step a) is mixed with astragalus powder at a mass ratio of (6-8):(2-4) to obtain a composite matrix; c) Pretreatment with ternary compound enzymatic hydrolysis Cellulase (50 U / g), pectinase (35 U / g), and β-glucanase (15 U / g) were added to the composite matrix, and hydrolysis was carried out at 48-52℃ and pH 4.8-5.2 for 1.5-2.5 hours. The enzymatic hydrolysis, combined with the gradient freezing treatment in step a), resulted in a matrix cell wall destruction rate of ≥99.1%. d) Substrate sterilization Add water to the enzymatically hydrolyzed matrix to a moisture content of 56%-59%, autoclave at 121℃ for 18-20 minutes, and cool to room temperature; e) Activation and compounding of tetrad strains Bifidobacterium longum was inoculated into MRS liquid medium and cultured anaerobically at 37°C until OD200. 600 =0.6-0.8, centrifuged and resuspended in sterile physiological saline to 1×10⁻⁶. 8 CFU / mL, β-glucosidase activity ≥112.3 U / g; Bacillus subtilis inoculated into LB liquid medium and cultured in a shaker at 37°C until OD... 600 =0.8-1.0, bacterial concentration 1×10 8 CFU / mL, protease activity ≥98.7 U / g; Saccharomyces cerevisiae inoculated into YPD liquid medium and cultured at 37℃ until OD 600 =0.6-0.8, bacterial concentration 1×10 8 CFU / mL, sucrase activity ≥135.2 U / g; *Lactobacillus plantarum* inoculated into MRS liquid medium and incubated statically at 37°C until OD500. 600 =0.7-0.9, bacterial concentration 1×10 8 CFU / mL, cAMP synthase activity ≥90.6U / g; four bacterial solutions were mixed in a volume ratio of (1.5-2.5):(1.5-2.5):(0.8-1.2):(0.8-1.2) and sprayed onto the substrate obtained in step d) at a total inoculum of 3%-4% (v / w); f) Adenosine concentration-responsive staged solid-state fermentation Under the condition of real-time monitoring of adenosine concentration in fermentation broth, the following parameters were controlled in stages: 0-10h: Temperature 38-39℃, pH 6.7-6.9, DO 6-10%. Maintain this stage when adenosine concentration ≤0.5mg / L to promote the proliferation of Bacillus subtilis and Lactobacillus plantarum. 10-36h: Temperature 36-37℃, pH 6.4-6.6, DO 0.5%-1%. When the adenosine concentration is ≥0.8mg / L, this stage is entered, which is the main fermentation stage, and the cAMP synthesis rate is ≥0.37μg / (g・h). 36-60h: Temperature 34-35℃, pH 6.2-6.4, DO < 0.3%, fermentation ends when the content of verbascoside isoflavone glucoside is ≥ 1.8 mg / g; g) Post-processing for product stabilization After fermentation, the mixture was freeze-dried at -50℃ until the moisture content was ≤5%, pulverized and passed through a 100-mesh sieve, and then microencapsulated with 10% (w / w) hydroxypropyl-β-cyclodextrin (inlet air temperature 180℃, outlet air temperature 80℃, spray rate 5mL / min) to obtain the fermented astragalus and jujube complex.

3. The method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function according to claim 2, characterized in that: In step a), the microwave pretreatment parameters were 2450MHz, 300W, and 1.5min. After gradient freezing and thawing, the cell destruction rate of jujube reached 98.7±1.2%, and the cell destruction rate of astragalus reached 96.5±1.5%. In step c), the hydrolysis conditions were 50℃ and pH 5.0 for 2 hours. After hydrolysis by the ternary complex enzyme, the cell wall destruction rate reached 99.3±0.8%.

4. The method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function according to claim 2, characterized in that: In step e), the inoculation volume ratio of the quaternary bacterial solution is 2:2:1:1, the total inoculation amount is 3.5% (v / w), and the total number of viable bacteria in the substrate after inoculation is ≥3×10⁻⁶. 7 CFU / g; The Lactobacillus plantarum is a Gram-positive facultative anaerobic bacterium with acid-resistant properties.

5. The method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function according to claim 1, characterized in that: The termination conditions for responsive fermentation in step f) are: cAMP content ≥ 13.0 μg / g and 6,8-diglucosyl-naringenin content ≥ 6.7 mg / g, at which point the total fermentation cycle is 48 hours.

6. The method for preparing a highly active fermented astragalus and jujube complex for improving myocardial function according to claim 1, characterized in that: In step g), the hydroxypropyl-β-cyclodextrin encapsulated in the microcapsules has a purity of ≥99%, and the encapsulated complex has a solubility of 18.5 mg / mL at 25 °C.

7. The fermented astragalus and jujube complex prepared by the method according to any one of claims 2-6, characterized in that: The complex contains cAMP content ≥13.0 μg / g (dry weight), total astragalus saponins content ≥9.5 mg / g, astragalus verbenae isoflavone glucoside content ≥1.9 mg / g, 6,8-digluxyl-2(S) and 2(R)-naringenin content in jujube ≥6.7 mg / g, cAMP degradation rate ≤2.3% after 3 months of storage at room temperature in the dark, and water solubility ≥18.5 mg / mL at 25℃.

8. The fermented astragalus and jujube complex according to claim 7, characterized in that: At a dose of 150 mg / kg, the compound reduced ST segment elevation by ≥53% and serum CK-MB activity by ≥39% in mice with myocardial ischemia (P<0.001); when combined with tanshinone extract at a mass ratio of 3:1, it inhibited hypoxia-induced cardiomyocyte apoptosis by ≥43.6%.

9. The application of the fermented astragalus and jujube complex according to claim 7 in the preparation of health food for myocardial ischemia pretreatment, characterized in that: The health food product is a chewable tablet or oral liquid, and each unit of the preparation contains 200-300mg of the complex, without the need for additional sweeteners, acidulants or preservatives.

10. The use of the fermented astragalus and jujube complex according to claim 7 in the preparation of a pharmaceutical composition for adjuvant treatment during the recovery period of myocardial infarction, characterized in that: The pharmaceutical composition comprises the complex and tanshinone extract in a mass ratio of 3:1, and can be prepared into tablets, capsules or granules for reducing cardiomyocyte apoptosis rate and improving cardiac function indicators.

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