A longevity-promoting American ginseng composite freeze-dried powder and its preparation method and application

Abstract

This invention belongs to the field of traditional Chinese medicine technology, and discloses a compound freeze-dried powder of American ginseng for longevity, its preparation method, and its application. The compound freeze-dried powder comprises, by weight, 25-35 parts of freeze-dried powder of American ginseng and 5-15 parts of freeze-dried powder of sea buckthorn, fermented to obtain freeze-dried powder A; 15-25 parts of Rhodiola rosea; 10-20 parts of freeze-dried powder of deer antler, enzymatically hydrolyzed to obtain freeze-dried powder B; 3-8 parts of Cornus officinalis; 2-5 parts of Allium macrostemon; 1-3 parts of Mentha haplocalyx; and 2-5 parts of Lophatherum gracile. This invention significantly enhances the efficacy of active ingredients through enzymatic hydrolysis and bacterial fermentation technology: American ginseng and sea buckthorn are mixed in a certain proportion and fermented successively with Lactobacillus bulgaricus CICC 6047 and Saccharomyces cerevisiae CICC1009; and deer antler is enzymatically hydrolyzed by Bacillus subtilis protease. The compound freeze-dried powder of the present invention has the effects of prolonging the life of nematodes and anti-oxidation; at the same time, it also exhibits a mild laxative effect. The laxative powder can shorten the time of first defecation in constipated mice and increase the number and weight of feces excreted, and is especially suitable for sub-healthy elderly people with long-term fatigue and constipation.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine technology, specifically relating to a compound freeze-dried powder of American ginseng for prolonging life, its preparation method and application. Background Technology

[0002] With the accelerating aging process and increasing awareness of health and wellness in modern society, the market demand for products with longevity-promoting effects is booming. Traditional Chinese medicine theory holds that aging is closely related to "deficiency of qi and blood, and imbalance of the five internal organs," and precious Chinese medicinal herbs such as American ginseng, rhodiola rosea, and deer antler have been recorded since ancient times as having the effects of "tonifying qi and blood, nourishing the liver and kidneys, and prolonging life." Modern scientific research has also confirmed that the active ingredients in these medicinal herbs, such as saponins, polysaccharides, and flavonoids, have significant antioxidant, immunomodulatory, and metabolic-improving effects, and can delay the aging process through multiple pathways. However, the active ingredients in these medicinal herbs often suffer from low bioavailability and poor stability, which seriously restricts their efficacy. Although existing technologies attempt to develop related products through simple compound formulations or conventional processing methods, it is difficult to overcome technical bottlenecks such as low absorption and utilization rates of active ingredients and poor targeting. In particular, for the complex physiological process of aging, traditional preparations cannot achieve multi-system synergistic regulation and cannot fully exert the longevity-promoting effects of medicinal herbs. Therefore, how to improve the bioavailability of active ingredients in medicinal materials through modern modification technology, achieve synergistic effects of multiple components, and develop innovative products with truly significant anti-aging functions has become a key issue that urgently needs to be addressed. Summary of the Invention

[0003] The purpose of this invention is to overcome the shortcomings of the prior art and provide a compound freeze-dried powder of American ginseng and its application in longevity and anti-aging products, so as to solve the problems mentioned in the above-mentioned technical background.

[0004] To achieve the above objectives, the present invention provides the following technical solution:

[0005] In a first aspect, the present invention provides a compound freeze-dried powder of American ginseng, comprising freeze-dried powder A obtained by fermentation after mixing 25-35 parts of freeze-dried American ginseng powder and 5-15 parts of freeze-dried sea buckthorn powder, 15-25 parts of Rhodiola rosea, freeze-dried powder B obtained by enzymatic hydrolysis of 10-20 parts of freeze-dried deer antler powder, 3-8 parts of Cornus officinalis, 2-5 parts of Allium macrostemon, 1-3 parts of Mentha haplocalyx, and 2-5 parts of Lophatherum gracile.

[0006] Preferably, the American ginseng compound freeze-dried powder contains the following components by weight: 30 parts American ginseng freeze-dried powder and 10 parts sea buckthorn freeze-dried powder mixed and fermented to obtain freeze-dried powder A, 20 parts Rhodiola rosea, 15 parts deer antler freeze-dried powder enzymatically hydrolyzed to obtain freeze-dried powder B, 5 parts Cornus officinalis, 3 parts Allium macrostemon, 2 parts Mentha haplocalyx, and 3 parts Lophatherum gracile.

[0007] Preferably, the freeze-dried American ginseng powder and freeze-dried sea buckthorn powder are mixed in a certain proportion and then fermented successively by brewer's yeast CICC1009 and Lactobacillus bulgaricus CICC6047.

[0008] Preferably, the freeze-dried deer antler powder is hydrolyzed by Bacillus subtilis protease.

[0009] Secondly, the present invention provides a method for preparing American ginseng compound freeze-dried powder, comprising the following steps:

[0010] (1) Preparation and fermentation of freeze-dried powder of American ginseng and sea buckthorn

[0011] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0012] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0013] 3) Vacuum drying: Place the product in a freeze-drying chamber, turn on the vacuum pump, and reduce the pressure inside the chamber to below 0.01 mbar. Slowly raise the temperature from -40℃ to 5℃ (heating rate approximately 1℃ / hour) for 30 hours; then gradually raise the temperature from 0℃ to 40℃ (heating rate 1℃ / hour) for 24 hours. After drying, pulverize the product using a pulverizer (≥200 mesh).

[0014] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water, remove the pits, and immediately pulp them using a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color-protecting agent to inhibit vitamin C oxidation, and then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the sea buckthorn pulp evenly into a 2-3mm thin layer to ensure uniform heat transfer, and place it in a -45℃ low temperature environment for quick freezing for 3 hours. Transfer the quick-frozen pulp to a freeze-drying chamber, control the vacuum degree ≤8Pa, and maintain the plate temperature at -25℃ for 20 hours. Gradually increase the temperature in stages: -5℃ (4h) → 15℃ (4h) → 30℃ (5h). Process the dried pulp blocks using a pulverizer (≥300 mesh).

[0015] 5) Fermentation: The fermentation system consists of 25-35 parts American ginseng freeze-dried powder, 5-15 parts sea buckthorn freeze-dried powder, 2.5 parts food-grade glucose, 0.01 parts magnesium sulfate, and 20 parts distilled water. A 5% (v / w) *Lactobacillus bulgaricus* CICC6047 bacterial solution (OD600 of the bacterial solution is 1.0-1.2) is inoculated. The mixture is allowed to ferment statically at 37℃ for 24 hours. After 24 hours, a 3% (v / w) *Saccharomyces cerevisiae* CICC1009 bacterial solution (OD600 of the bacterial solution is 1.0-1.2) is inoculated. Fermentation is then carried out at 30℃ with stirring for 48 hours, producing a slight alcoholic aroma. After fermentation, *Lactobacillus bulgaricus* CICC6047 and *Saccharomyces cerevisiae* CICC1009 in the fermented product are inactivated using a 3% H2O2 solution. The inoculation amount is based on the total mass of the American ginseng freeze-dried powder and sea buckthorn freeze-dried powder.

[0016] 6) Powdering: The fermented material is filtered to obtain the filtrate. The filtrate is concentrated under reduced pressure at 50℃ to 1 / 5 of its original volume, and then 5% maltodextrin is added for pre-freezing treatment (4℃→-20℃→-40℃, 8 hours). Subsequently, it is freeze-dried in two stages: the first stage is -35℃→0℃ (24 hours) to remove free water, and the second stage is 0℃→25℃ (8 hours) to remove bound water. After freeze-drying, the material is pulverized at low temperature to 200 mesh.

[0017] (2) Preparation and enzymatic hydrolysis of freeze-dried deer antler powder

[0018] 1) Cleaning: Thaw the frozen fresh antler tip tissue and slice it. Clean the antler tip with pre-cooled distilled water at 4°C.

[0019] Cut the tissue into thin slices until the bloodstains are washed away, and then cut them into slices about 5mm thick.

[0020] 2) Quick-freezing: Spread the sliced ​​deer antlers flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0021] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 0℃ for 30 hours; gradually raise the temperature from 0℃ to 30℃ for 16 hours, and then pulverize with a pulverizer (≥300 mesh).

[0022] 4) Enzymatic hydrolysis: Weigh out Bacillus subtilis protease and dissolve it in PBS solution at pH 7.1. After complete dissolution, add the freeze-dried deer antler powder for enzymatic hydrolysis. The hydrolysis temperature is 52℃, the pH value is 7.1, and the time is 3 hours. The enzyme activity of Bacillus subtilis protease is 150 U / mg, the amount of enzyme added is 0.05% of the freeze-dried deer antler powder, and the mass fraction of freeze-dried deer antler powder is 15% (w / v, the proportion in the enzymatic hydrolysis system).

[0023] pH 7.1 PBS solution: Take 33 ml of 0.2 M NaH2PO4 solution and 67 ml of 0.2 M Na2HPO4 solution, mix them well to obtain pH 7.1 PBS buffer.

[0024] 5) Powdering: After pre-freezing the supernatant after enzymatic hydrolysis at -40℃, it is dried in stages under vacuum of 25 Pa (sublimation at -20℃, desorption at 27℃), and finally ground to obtain freeze-dried powder (≥200 mesh).

[0025] (3) Rhodiola rosea freeze-dried powder

[0026] The pulverized Rhodiola rosea (100 mesh) was placed in water (1g dissolved in 10ml of water) and extracted at 85℃ for 2 hours. After filtration, it was concentrated to 1 / 5 of its volume, pre-frozen at -40℃ for 4 hours, and then freeze-dried in stages under 13Pa vacuum (-25℃ for 12h → 30℃ for 6h), and finally ultra-finely pulverized to 200 mesh.

[0027] (4) Cornus officinalis

[0028] After pulverizing the dried Cornus officinalis to 30 mesh, it was placed in water (1g dissolved in 10ml of water) and extracted at 80℃ for 2 hours. After filtration, it was concentrated to 1 / 5 of its volume. After pre-freezing at -40℃ for 4 hours, it was freeze-dried in stages under 15Pa vacuum (-25℃ for 12h → 30℃ for 6h) and finally pulverized to 200 mesh.

[0029] (5) Xie Bai

[0030] Fresh bulbs were selected, and the outer fibrous membrane was manually peeled off. The bulbs were then cleaned with ultrasound (40kHz) and soaked in 1% nisin solution for 5 minutes to inhibit spores. After treatment, the bulbs were longitudinally cut into 3mm thin slices. The slices were then placed in a 0.3% cysteine ​​+ 1% salt color-protecting solution and frozen at -50℃ for 4 hours. After freezing, the slices were placed in a freeze-drying chamber with a vacuum degree ≤5Pa and maintained at -30℃ for 22 hours. Then, the bulbs were dried in stages: -10℃ (2h) → 0℃ (3h) → 25℃ (4h). The resulting bulb blocks were then subjected to air-jet pulverization (≥300 mesh) at 5℃.

[0031] (6) Peppermint

[0032] Fresh mint was selected, and impurities were removed by ultrasonic cleaning (40kHz / 3min). It was then disinfected with a compound of 0.05% sodium hypochlorite and 0.1% citric acid. After rinsing, it was centrifuged and dehydrated. The leaves were then crushed into 2-3mm fragments at low temperature (4℃), and the stems were micro-shorn into 1cm segments. Immediately after cutting, the stems were immersed in a compound color-protecting solution of 0.2% ascorbic acid and 0.5% trehalose. After being laid out, the stems were quickly frozen at -50℃ for 2 hours. After quick-freezing, the stems were placed in a freeze-drying chamber with a vacuum degree ≤5Pa and maintained at -35℃ for 18 hours. Finally, the temperature was gradually increased: -20℃ (3h) → 0℃ (3h) → 25℃ (3h). Finally, the stems were pulverized at low temperature (-15℃) (≥300 mesh).

[0033] (7) Lophatherum gracile

[0034] Fresh bamboo leaves were selected, the surface wax layer was removed, and then soaked in 0.1% sodium bicarbonate solution for 2 minutes. After being quick-frozen with liquid nitrogen, they were brittled and broken into 2-3 mm fragments (-196℃ / 1min). After being laid flat, they were quick-frozen at -45℃ for 3 hours, placed in a freeze-drying chamber, with a vacuum degree ≤10Pa, and the plate temperature maintained at -30℃ for 16 hours. Then, the temperature was gradually increased: -15℃ (3h) → 5℃ (4h) → 25℃ (5h), and then crushed in a pulverizer (≥400 mesh).

[0035] (8) Compound

[0036] The freeze-dried powder obtained in steps (1)-(7) is compounded with 25-35 parts of American ginseng freeze-dried powder and 5-15 parts of sea buckthorn freeze-dried powder and fermented to obtain freeze-dried powder A, 15-25 parts of Rhodiola rosea, 10-20 parts of deer antler freeze-dried powder B obtained by enzymatic hydrolysis, 3-8 parts of Cornus officinalis, 2-5 parts of Allium macrostemon, 1-3 parts of peppermint, and 2-5 parts of Lophatherum gracile to obtain the final product.

[0037] Secondly, the present invention provides the aforementioned American ginseng compound freeze-dried powder and its application in longevity and anti-aging products.

[0038] Compared with the prior art, the beneficial effects of the present invention are:

[0039] 1. In this application, brewer's yeast CICC1009 and Lactobacillus bulgaricus CICC6047 were used to ferment American ginseng and sea buckthorn. Bacillus subtilis protease was used to enzymatically hydrolyze deer antler to improve the yield of deer antler polypeptides. The fermentation and hydrolysis products were then compounded with Rhodiola rosea, Cornus officinalis, Allium macrostemon, Mentha haplocalyx, and Lophatherum gracile to form a freeze-dried powder. The synergistic effect among the components in the American ginseng compound freeze-dried powder significantly prolongs the lifespan of nematodes and scavenge DPPH free radicals.

[0040] 2. The freeze-dried powder of this application, the fermented products of American ginseng and sea buckthorn, and menthol work synergistically to stimulate intestinal peristalsis, increase stool volume, and lubricate the intestines, thus giving it a mild laxative effect.

[0041] 3. The freeze-dried powder disclosed in this application can prolong the lifespan of nematodes, exhibits excellent antioxidant effects in in vitro experiments, and has a very prominent anti-aging effect, showing great market application prospects. Attached Figure Description

[0042] Figure 1 The figure shows the experimental results of the effect of the American ginseng compound freeze-dried powder of the present invention on the lifespan of nematodes.

[0043] Figure 2 The figure shows the experimental results of the effect of the American ginseng compound freeze-dried powder of the present invention on the DPPH free radical scavenging rate;

[0044] Figure 3 The figure shows the experimental results of the effect of the American ginseng compound freeze-dried powder of the present invention on the defecation of mice. Detailed Implementation

[0045] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that, unless otherwise specified, the following embodiments and features described therein can be combined with each other.

[0046] Some of the raw materials used in this application:

[0047] Rhodiola rosea and Cornus officinalis are dried products, and their quality meets the 2020 Pharmacopoeia standards.

[0048] American ginseng, deer antler, sea buckthorn, allium macrostemon, mint, and bamboo leaves are all fresh and were purchased from Huinong.com.

[0049] Lactobacillus bulgaricus CICC6047 was purchased from the China Industrial Microbial Culture Collection Center.

[0050] Saccharomyces cerevisiae CICC1009 was purchased from the China Industrial Microbial Culture Collection Center.

[0051] Saccharomyces cerevisiae CICC1001 was purchased from the China Industrial Microbial Culture Collection Center.

[0052] Bacillus subtilis protease, specification 150U / mg, product number: 9014-01-1, purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0053] Flavor protease, specification 20U / mg, product number: S10153, purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0054] Example 1

[0055] A compound freeze-dried powder of American ginseng, the formula of which includes: freeze-dried powder A obtained by fermentation after mixing 30 parts of freeze-dried American ginseng powder and 10 parts of freeze-dried sea buckthorn powder, 20 parts of Rhodiola rosea, freeze-dried powder B obtained by enzymatic hydrolysis of freeze-dried deer antler powder, 5 parts of Cornus officinalis, 3 parts of Allium macrostemon, 2 parts of Mentha haplocalyx, and 3 parts of Lophatherum gracile.

[0056] The freeze-dried American ginseng powder and freeze-dried sea buckthorn powder were mixed in a certain proportion and then fermented with Lactobacillus bulgaricus CICC6047 and Saccharomyces cerevisiae CICC1009.

[0057] The freeze-dried deer antler powder mentioned above is treated with Bacillus subtilis protease.

[0058] Example 2

[0059] A compound freeze-dried powder of American ginseng, the formula of which includes: freeze-dried powder A obtained by fermentation after mixing 25 parts of freeze-dried American ginseng powder and 15 parts of freeze-dried sea buckthorn powder, 25 parts of Rhodiola rosea, freeze-dried powder B obtained by enzymatic hydrolysis of 10 parts of freeze-dried deer antler powder, 3 parts of Cornus officinalis, 2 parts of Allium macrostemon, 3 parts of Mentha haplocalyx, and 2 parts of Lophatherum gracile.

[0060] The preparation method of the American ginseng compound freeze-dried powder in this embodiment is the same as that in Example 1.

[0061] Example 3

[0062] A compound freeze-dried powder of American ginseng, the formula of which, by weight percentage, includes: 35 parts freeze-dried powder of American ginseng and 5 parts freeze-dried powder of sea buckthorn mixed and fermented to obtain freeze-dried powder A, 15 parts Rhodiola rosea, 20 parts freeze-dried powder of deer antler obtained by enzymatic hydrolysis to obtain freeze-dried powder B, 8 parts Cornus officinalis, 5 parts Allium macrostemon, 1 part Mentha haplocalyx and 5 parts Lophatherum gracile.

[0063] The preparation method of the American ginseng compound freeze-dried powder in this embodiment is the same as that in Example 1.

[0064] Comparative Example 1

[0065] This comparative example is similar to Example 1, except that the American ginseng and sea buckthorn were only fermented with Lactobacillus bulgaricus CICC6047.

[0066] Comparative Example 2

[0067] This comparative example is similar to Example 1, except that: sea buckthorn and sea buckthorn were only fermented with brewer's yeast CICC1009.

[0068] Comparative Example 3

[0069] This comparative example is similar to Example 1, except that neither American ginseng nor sea buckthorn is fermented.

[0070] Comparative Example 4

[0071] This comparative example is similar to Example 1, except that the American ginseng and sea buckthorn are fermented with Saccharomyces cerevisiae CICC1009 and Lactobacillus bulgaricus CICC6047 instead of Saccharomyces cerevisiae CICC1001 and Lactobacillus bulgaricus CICC6047.

[0072] Comparative Example 5

[0073] This comparative example is similar to Example 1, except that the deer antler is not hydrolyzed by Bacillus subtilis protease, but by flavor protease.

[0074] Comparative Example 6

[0075] This comparative example is similar to Example 1, except that the deer antler is not enzymatically hydrolyzed but directly made into freeze-dried powder and then compounded.

[0076] Comparative Example 7

[0077] This comparative example is similar to Example 1, except that American ginseng was omitted.

[0078] Comparative Example 8

[0079] This comparative example is similar to Example 1, except that sea buckthorn is removed.

[0080] Comparative Example 9

[0081] This comparative example is similar to Example 1, except that Rhodiola rosea is omitted.

[0082] Comparative Example 10

[0083] This comparative example is similar to Example 1, except that the deer antler is removed.

[0084] Comparative Example 11

[0085] This comparative example is similar to Example 1, except that mint was removed.

[0086] The American ginseng compound freeze-dried powder in Example 1 was prepared according to the following method:

[0087] (1) Preparation and fermentation of freeze-dried powder of American ginseng and sea buckthorn

[0088] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0089] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0090] 3) Vacuum drying: Place the product in a freeze-drying chamber, turn on the vacuum pump, and reduce the pressure inside the chamber to below 0.01 mbar. Slowly raise the temperature from -40℃ to 5℃ (heating rate approximately 1℃ / hour) for 30 hours; then gradually raise the temperature from 0℃ to 40℃ (heating rate 1℃ / hour) for 24 hours. After drying, pulverize the product to 200 mesh using a pulverizer.

[0091] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water, remove the pits, and immediately pulp them using a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color-protecting agent to inhibit vitamin C oxidation, and then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the sea buckthorn pulp evenly into a 2-3mm thin layer to ensure uniform heat transfer, and place it in a -45℃ low temperature environment for quick freezing for 3 hours. Transfer the quick-frozen pulp to a freeze-drying chamber, control the vacuum degree ≤8Pa, and maintain the plate temperature at -25℃ for 20 hours. Gradually increase the temperature in stages: -5℃ (4h) → 15℃ (4h) → 30℃ (5h). Process the dried pulp blocks to 300 mesh using a pulverizer.

[0092] 5) Fermentation: The fermentation system consisted of 30 parts American ginseng freeze-dried powder, 10 parts sea buckthorn freeze-dried powder, 2.5 parts food-grade glucose, 0.01 parts magnesium sulfate, and 20 parts distilled water. A 5% (v / w) *Lactobacillus bulgaricus* CICC6047 bacterial solution (OD600 of the bacterial solution was 1.0) was inoculated, and the mixture was allowed to ferment statically at 37°C for 24 hours. Then, a 3% (v / w) *Saccharomyces cerevisiae* CICC1009 bacterial solution (OD600 of the bacterial solution was 1.0) was inoculated, and the mixture was stirred and fermented at 30°C for 48 hours, producing a slight alcoholic aroma. After fermentation, *Lactobacillus bulgaricus* CICC6047 and *Saccharomyces cerevisiae* CICC1009 in the fermented product were inactivated using a 3% H2O2 solution. The inoculation amount was based on the total mass of the American ginseng freeze-dried powder and sea buckthorn freeze-dried powder.

[0093] 6) Powdering: The fermented material is filtered to obtain the filtrate. The filtrate is concentrated under reduced pressure at 50℃ to 1 / 5 of its original volume, and then 5% maltodextrin is added for pre-freezing treatment (4℃→-20℃→-40℃, 8 hours). Subsequently, it is freeze-dried in two stages: the first stage is -35℃→0℃ (24 hours) to remove free water, and the second stage is 0℃→25℃ (8 hours) to remove bound water. After freeze-drying, the material is pulverized at low temperature to 200 mesh.

[0094] (2) Preparation and enzymatic hydrolysis of freeze-dried deer antler powder

[0095] 1) Cleaning: Thaw the frozen fresh antler tip tissue and slice it. Clean the antler tip with pre-cooled distilled water at 4°C.

[0096] Cut the tissue into thin slices until the bloodstains are washed away, and then cut them into slices about 5mm thick.

[0097] 2) Quick-freezing: Spread the sliced ​​deer antlers flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0098] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 0℃ for 30 hours; gradually raise the temperature from 0℃ to 30℃ for 16 hours, and pulverize to 300 mesh using a pulverizer.

[0099] 4) Enzymatic hydrolysis: Weigh out Bacillus subtilis protease and dissolve it in PBS solution at pH 7.1. After dissolution, add the freeze-dried deer antler powder for enzymatic hydrolysis. The hydrolysis temperature is 52℃, the pH value is 7.1, and the time is 3 hours. The enzyme activity of Bacillus subtilis protease is 150 U / mg, the amount of enzyme added is 0.05% of the freeze-dried deer antler powder, and the mass fraction of freeze-dried deer antler powder is 15% (w / v, the proportion in the enzymatic hydrolysis system).

[0100] 5) Powdering: After pre-freezing the supernatant after enzymatic hydrolysis at -40℃, it is dried in stages under vacuum of 25 Pa (sublimation at -20℃, desorption at 27℃), and finally ground to obtain freeze-dried powder to 200 mesh.

[0101] (3) Rhodiola rosea freeze-dried powder

[0102] The pulverized Rhodiola rosea (100 mesh) was placed in water (1g dissolved in 10ml of water) and extracted at 85℃ for 2 hours. After filtration, it was concentrated to 1 / 5 of its volume, pre-frozen at -40℃ for 4 hours, and then freeze-dried in stages under 13Pa vacuum (-25℃ for 12h → 30℃ for 6h), and finally ultra-finely pulverized to 200 mesh.

[0103] (4) Cornus officinalis

[0104] After pulverizing the dried Cornus officinalis to 30 mesh, it was placed in water (1g dissolved in 10ml of water) and extracted at 80℃ for 2 hours. After filtration, it was concentrated to 1 / 5 of its volume. After pre-freezing at -40℃ for 4 hours, it was freeze-dried in stages under 15Pa vacuum (-25℃ for 12h → 30℃ for 6h) and finally ultra-fine pulverized to 200 mesh.

[0105] (5) Xie Bai

[0106] Fresh bulbs were selected, and the outer fibrous membrane was manually peeled off. The bulbs were then cleaned with ultrasound (40kHz) and soaked in 1% nisin solution for 5 minutes to inhibit spores. After treatment, the bulbs were longitudinally cut into 3mm thin slices. The slices were then placed in a 0.3% cysteine ​​+ 1% salt color-protecting solution and frozen at -50℃ for 4 hours. After freezing, the slices were placed in a freeze-drying chamber with a vacuum degree ≤5Pa and maintained at -30℃ for 22 hours. Then, the bulbs were dried in stages: -10℃ (2h) → 0℃ (3h) → 25℃ (4h). The resulting bulb blocks were then pulverized to 300 mesh at 5℃ using airflow.

[0107] (6) Peppermint

[0108] Fresh mint was selected, and impurities were removed by ultrasonic cleaning (40kHz / 3min). It was then disinfected with a compound of 0.05% sodium hypochlorite and 0.1% citric acid. After rinsing, it was centrifuged and dehydrated. The leaves were then crushed into 2-3mm fragments at low temperature (4℃), and the stems were micro-sheared into 1cm segments. Immediately after shearing, the stems were immersed in a compound color-protecting solution of 0.2% ascorbic acid and 0.5% trehalose. After being laid out, the stems were quickly frozen at -50℃ for 2 hours. After quick-freezing, the stems were placed in a freeze-drying chamber with a vacuum degree ≤5Pa and maintained at -35℃ for 18 hours. Finally, the temperature was gradually increased: -20℃ (3h) → 0℃ (3h) → 25℃ (3h). Finally, the stems were pulverized to 300 mesh at low temperature (-15℃) using a pulverizer.

[0109] (7) Lophatherum gracile

[0110] Fresh bamboo leaves were selected, the surface wax layer was removed, and then soaked in 0.1% sodium bicarbonate solution for 2 minutes. After being quick-frozen with liquid nitrogen, they were brittled and broken into 2-3 mm fragments (-196℃ / 1min). After being laid flat, they were quick-frozen at -45℃ for 3 hours, placed in a freeze-drying chamber, with a vacuum degree ≤10Pa, and the plate temperature maintained at -30℃ for 16 hours. Then, the temperature was gradually increased: -15℃ (3h) → 5℃ (4h) → 25℃ (5h), and then pulverized in a pulverizer to 400 mesh.

[0111] (8) Compound

[0112] The freeze-dried powder obtained in steps (1)-(7) is compounded with freeze-dried powder A obtained by fermentation after mixing 30 parts of American ginseng freeze-dried powder and 10 parts of sea buckthorn freeze-dried powder, 20 parts of Rhodiola rosea, 15 parts of deer antler freeze-dried powder obtained by enzymatic hydrolysis, 5 parts of Cornus officinalis freeze-dried powder, 3 parts of Allium macrostemon freeze-dried powder, 2 parts of peppermint freeze-dried powder, and 3 parts of Lophatherum gracile freeze-dried powder to obtain the final product.

[0113] Preparation method of Comparative Example 1:

[0114] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0115] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0116] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 5℃ (heating rate of about 1℃ / hour) for 30 hours; gradually raise the temperature from 0℃ to 40℃ (heating rate of 1℃ / hour) for 24 hours, and then pulverize to 200 mesh using a pulverizer.

[0117] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water, remove the pits, and immediately pulp them using a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color-protecting agent to inhibit vitamin C oxidation, and then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the sea buckthorn pulp evenly into a 2-3mm thin layer to ensure uniform heat transfer, and place it in a -45℃ low temperature environment for quick freezing for 3 hours. Transfer the quick-frozen pulp to a freeze-drying chamber, control the vacuum degree ≤8Pa, and maintain the plate temperature at -25℃ for 20 hours. Gradually increase the temperature in stages: -5℃ (4h) → 15℃ (4h) → 30℃ (5h). Process the dried pulp blocks to 300 mesh using a pulverizer.

[0118] 5) Fermentation: The fermentation system consisted of 30 parts American ginseng freeze-dried powder, 10 parts sea buckthorn freeze-dried powder, 2.5 parts food-grade glucose, 0.01 parts magnesium sulfate, and 20 parts distilled water. 5% *Lactobacillus bulgaricus* CICC6047 (OD600 = 1.0) was inoculated, and the mixture was allowed to ferment at 37°C for 24 hours. After fermentation, the *Lactobacillus bulgaricus* CICC6047 in the fermentation product was inactivated using a 3% H2O2 solution.

[0119] 6) Powdering: The fermented material is filtered to obtain the filtrate. The filtrate is concentrated under reduced pressure at 50℃ to 1 / 5 of its original volume, and then 5% maltodextrin is added for pre-freezing treatment (4℃→-20℃→-40℃, 8 hours). Subsequently, it is freeze-dried in two stages: the first stage is -35℃→0℃ (24 hours) to remove free water, and the second stage is 0℃→25℃ (8 hours) to remove bound water. After freeze-drying, the material is pulverized at low temperature to 200 mesh.

[0120] The remaining steps are the same as the preparation method in Example 1.

[0121] Preparation method of Comparative Example 2:

[0122] Preparation and fermentation of freeze-dried American ginseng and sea buckthorn powder

[0123] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0124] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0125] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 5℃ (heating rate of about 1℃ / hour) for 30 hours; gradually raise the temperature from 0℃ to 40℃ (heating rate of 1℃ / hour) for 24 hours, and then pulverize to 200 mesh using a pulverizer.

[0126] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water, remove the pits, and immediately pulp them with a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color protectant to inhibit vitamin C oxidation, and then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the sea buckthorn pulp evenly into a 2-3mm thin layer to ensure uniform heat transfer, and place it in a -45℃ low temperature environment for quick freezing for 3 hours. Transfer the quick-frozen pulp to a freeze-drying chamber, control the vacuum degree ≤8Pa, and maintain the plate temperature at -25℃ for 20 hours. Gradually increase the temperature in stages: -5℃ (4h) → 15℃ (4h) → 30℃ (5h). Process the dried pulp blocks to 300 mesh using a pulverizer.

[0127] 5) Fermentation: The fermentation system consisted of 30 parts freeze-dried American ginseng powder, 10 parts freeze-dried sea buckthorn powder, 2.5 parts food-grade glucose, 0.01 parts magnesium sulfate, and 20 parts distilled water. 3% Saccharomyces cerevisiae CICC1009 (OD600 1.0) was added, and fermentation was carried out at 30℃ with stirring for 48 hours, producing a slight alcoholic aroma. After fermentation, the Saccharomyces cerevisiae CICC1009 in the fermented product was inactivated using a 3% H2O2 solution.

[0128] 6) Powdering: The fermented material is filtered to obtain the filtrate. The filtrate is concentrated under reduced pressure at 50℃ to 1 / 5 of its original volume, and then 5% maltodextrin is added for pre-freezing treatment (4℃→-20℃→-40℃, 8 hours). Subsequently, it is freeze-dried in two stages: the first stage is -35℃→0℃ (24 hours) to remove free water, and the second stage is 0℃→25℃ (8 hours) to remove bound water. After freeze-drying, the material is pulverized at low temperature to 200 mesh.

[0129] The remaining steps are the same as the preparation method in Example 1.

[0130] Preparation method of Comparative Example 3:

[0131] Preparation of freeze-dried powders of American ginseng and sea buckthorn

[0132] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0133] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0134] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 5℃ (heating rate of about 1℃ / hour) for 30 hours; gradually raise the temperature from 0℃ to 40℃ (heating rate of 1℃ / hour) for 24 hours, and then pulverize to 200 mesh using a pulverizer.

[0135] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water. After washing, remove the pits, and immediately pulp them using a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color-protecting agent to inhibit vitamin C oxidation. Then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the obtained sea buckthorn pulp into a 3-5mm thin layer and quick-freeze at -50℃ for 2 hours. After quick-freezing, place it in a freeze-drying chamber with a vacuum degree ≤10Pa and a plate temperature of -30℃ for 24 hours. Then perform a stepwise temperature increase: -10℃ (4h) → 10℃ (4h) → 30℃ (6h). The freeze-dried sea buckthorn blocks are crushed to 200 mesh using a pulverizer. Mix 30 parts of American ginseng freeze-dried powder and 10 parts of sea buckthorn freeze-dried powder to obtain the final product.

[0136] The remaining steps are the same as the preparation method in Example 1.

[0137] Preparation method of Comparative Example 4:

[0138] Preparation and fermentation of freeze-dried American ginseng and sea buckthorn powder

[0139] 1) Cleaning: Rinse the surface of mud and sand with a soft brush and running water to avoid damaging the skin, and cut into thin slices of 2-3mm.

[0140] 2) Quick-freezing: Spread the American ginseng slices flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0141] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 5℃ (heating rate of about 1℃ / hour), and continue for 24-36 hours; gradually raise the temperature from 0℃ to 40℃ (heating rate of 1℃ / hour), and continue for 24 hours. After drying, pulverize to 200 mesh using a pulverizer.

[0142] 4) Select fresh sea buckthorn fruits with a maturity of ≥90%, soak them in a 1% sodium bicarbonate solution for 3 minutes to remove the surface wax, then wash them with running water, remove the pits, and immediately pulp them using a pulping machine. Add 0.1% vitamin E and 0.5% citric acid compound color-protecting agent to inhibit vitamin C oxidation, and then pasteurize (65℃, 30s) to obtain sea buckthorn pulp. Spread the obtained sea buckthorn pulp into a 3-5mm thin layer and quick-freeze at -50℃ for 2 hours. After quick-freezing, place it in a freeze-drying chamber with a vacuum degree ≤10Pa and a plate temperature of -30℃ for 24 hours. Then, perform a stepwise temperature increase: -10℃ (4h) → 10℃ (4h) → 30℃ (6h). The freeze-dried sea buckthorn blocks are crushed to 200 mesh using a pulverizer.

[0143] 5) Fermentation: The fermentation system consisted of 30 parts freeze-dried American ginseng powder, 10 parts freeze-dried sea buckthorn powder, 2.5 parts food-grade glucose, 0.01 parts magnesium sulfate, and 20 parts distilled water. 5% *Lactobacillus bulgaricus* CICC6047 (OD600 = 1.0) was inoculated, and the mixture was allowed to ferment statically at 37°C for 24 hours. Then, 3% *Saccharomyces cerevisiae* CICC1001 (OD600 = 1.0) was inoculated, and the mixture was stirred and fermented at 30°C for 48 hours, producing a slight alcoholic aroma. After fermentation, *Lactobacillus bulgaricus* CICC6047 and CICC1001 in the fermented product were inactivated using a 3% H2O2 solution.

[0144] 6) Powdering: The fermented material is filtered to obtain the filtrate. The filtrate is concentrated under reduced pressure at 50℃ to 1 / 5 of its original volume, and then 5% maltodextrin is added for pre-freezing treatment (4℃→-20℃→-40℃, 8 hours). Subsequently, it is freeze-dried in two stages: the first stage is -35℃→0℃ (24 hours) to remove free water, and the second stage is 0℃→25℃ (8 hours) to remove bound water. After freeze-drying, the material is pulverized at low temperature to 200 mesh.

[0145] The preparation steps for the remaining raw materials are the same as those in Example 1.

[0146] Preparation method of Comparative Example 5:

[0147] Preparation and enzymatic hydrolysis of freeze-dried deer antler powder

[0148] 1) Cleaning: Thaw the frozen fresh antler tip tissue and slice it. Use pre-cooled distilled water at 4°C to wash the antler tip tissue slices until the bloodstains are removed. Cut into slices about 5mm thick.

[0149] 2) Quick-freezing: Spread the sliced ​​deer antlers flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0150] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 0℃ for 30 hours; gradually raise the temperature from 0℃ to 30℃ for 16 hours, and pulverize to 300 mesh using a pulverizer.

[0151] 4) Enzymatic hydrolysis: Weigh the flavor protease and dissolve it in PBS solution at pH 7.1. After complete dissolution, add the freeze-dried deer antler powder for enzymatic hydrolysis. The hydrolysis temperature is 47℃, the pH value is 7.1, and the time is 3 hours. The enzyme activity of the flavor protease is 20 U / mg, the amount of enzyme added is 0.375% of the freeze-dried deer antler powder, and the mass fraction of the freeze-dried deer antler powder is 15% (w / v, the proportion in the enzymatic hydrolysis system).

[0152] 5) Powdering: After pre-freezing the supernatant after enzymatic hydrolysis at -40℃, it is dried in stages under vacuum of 25 Pa (sublimation at -20℃, desorption at 27℃), and finally ground to obtain freeze-dried powder to 200 mesh.

[0153] The preparation steps for the remaining raw materials are the same as those in Example 1.

[0154] Preparation method of Comparative Example 6:

[0155] Preparation of freeze-dried deer antler powder

[0156] 1) Cleaning: Thaw the frozen fresh antler tip tissue and slice it. Clean the antler tip with pre-cooled distilled water at 4°C.

[0157] Cut the tissue into thin slices until the bloodstains are washed away, and then cut them into slices about 5mm thick.

[0158] 2) Quick-freezing: Spread the sliced ​​deer antlers flat on a freeze-drying tray with a thickness of ≤1cm, and place them in a -45℃ ultra-low temperature freezer for 4 hours.

[0159] 3) Vacuum drying: Place in the freeze-drying chamber, turn on the vacuum pump, reduce the pressure inside the chamber to below 0.01 mbar, slowly raise the temperature from -40℃ to 0℃ for 30 hours; gradually raise the temperature from 0℃ to 30℃ for 16 hours, and pulverize to 200 mesh using a pulverizer.

[0160] To further clarify the efficacy of the American ginseng compound freeze-dried powder provided in the embodiments of this application, the following performance tests were also conducted in the embodiments of this application:

[0161] 1. Experiments to extend the lifespan of nematodes

[0162] Escherichia coli OP50, product number: HZB452999, purchased from Gray Algae Biotechnology Co., Ltd.

[0163] Caenorhabditis elegans, product number: X2344, purchased from Baosai Biotechnology Co., Ltd.

[0164] Reagent preparation:

[0165] Nematode growth medium (NGM): Add 0.25g tryptone, 0.3g sodium chloride and 2.1g agar powder to 100mL distilled water, sterilize at 121℃ for 20min. After the medium cools to a temperature that is not too hot to touch, add 2.5mL K2HPO4-KOH buffer (1 mol / L, pH 6.0), 0.1mL cholesterol solution (5 mg / mL, dissolved in 95% ethanol), 0.1mL MgSO4 solution (1 mol / L), and 0.1mL CaCl2 solution (1 mol / L). Shake well, pour the mixture onto a plate, and after solidification, store at 4℃ for later use.

[0166] M9 buffer: Add 6g Na2HPO4, 3g KH2PO4, 5g NaCl and 0.25g MgSO4•7H2O to 1L of distilled water, filter out the residue, and sterilize at 121℃ for 20min.

[0167] LB medium: Add 0.5g NaCl, 0.5g tryptone and 0.25g yeast extract to 50mL distilled water, and sterilize at 121℃ for 15min.

[0168] Nematode lysate: Add 0.1g NaOH and 0.5mL NaClO2 to 9.5mL distilled water, and prepare fresh before use.

[0169] Specific experimental methods:

[0170] Obtaining L4 stage nematodes: This invention employs a lysis synchronization method. 100 μL of E. coli OP50 (OD600=1.0) is pipetted onto the surface of NGM medium, gently spread, dried in a clean bench, and incubated overnight at 37°C in an inverted incubator. The nematodes are then transferred to NGM medium. Synchronization can be performed when a large number of nematodes are in the oviposition stage on the medium. The nematodes are washed off with M9 solution three times for sterilization, centrifuged at 200 rpm, and the supernatant is discarded. 1 mL of hypochlorite lysis buffer (nematode lysis buffer) is added, and the mixture is vortexed for 3 min. After centrifugation at 5000 rpm, the supernatant is discarded. Another 1 mL of lysis buffer is added, and the mixture is vortexed for 2 min. The EP tube is observed under a microscope until complete necrosis of the nematodes is achieved. The tube is then centrifuged at 5000 rpm to remove the supernatant. Wash the eggs with M9 medium and centrifuge at the same speed approximately 2-3 times until there is no pungent odor. Place the eggs on unstrained NGM medium and incubate for 14 hours to obtain L1 stage nematodes. Transfer the nematodes to sterilized medium and incubate for 2 days to obtain L4 stage nematodes, which can be used for experiments.

[0171] The American ginseng compound freeze-dried powders from Examples 1 and Comparative Examples 1-10 were prepared to a concentration of 50 μg / mL using distilled water. 100 μL of distilled water and 100 μL of the compound solution were mixed with 100 μL of *E. coli* OP50 (OD600=1.0) and evenly spread onto the surface of nematode growth medium (NGM) to prepare NGM plates with different compositions. A control group was also prepared by evenly spreading a mixture of distilled water and *E. coli* OP50 onto the surface of the nematode culture medium. Synchronized L4 stage *C. elegans* N2 were picked and added to the aforementioned NGM plates containing different compositions, with 45 nematodes per plate. The plates were incubated at 20°C, and this was recorded as day 0. The number of surviving, dead, and accidental deaths was recorded daily until all nematodes had died. The criteria for death were: no reaction when the nematodes were lightly touched with a needle, no swallowing, and the removal of nematodes that were burrowed into the agar or dried up on the plate wall.

[0172] according to Figure 1 As shown, the lifespan of nematodes in Example 1 was higher than that in the control group. Compared with Example 1, the results of Comparative Examples 1, 2, and 3 indicate that American ginseng and sea buckthorn need to be mixed and fermented to exert their life-extending effect; the results of Comparative Example 4 indicate that American ginseng and sea buckthorn need to be fermented with specific strains, namely Saccharomyces cerevisiae CICC1009 and Lactobacillus bulgaricus CICC6047, to have a life-extending effect; the results of Comparative Examples 5 and 6 indicate that deer antler, without enzymatic hydrolysis by Bacillus subtilis protease, had a much lower life-extending effect than Example 1; the results of Comparative Examples 1-10 indicate that fermenting American ginseng and sea buckthorn with Saccharomyces cerevisiae CICC1009 and Lactobacillus bulgaricus CICC6047, and enzymatically hydrolyzing deer antler with Bacillus subtilis protease to improve the yield of deer antler polypeptides, and then compounding the fermentation and hydrolysis products with Rhodiola rosea to make freeze-dried powder, the synergistic effect of the components in the freeze-dried powder composition has a significant effect on extending the lifespan of nematodes.

[0173] 2. DPPH free radical scavenging test

[0174] Specific experimental methods:

[0175] The American ginseng compound freeze-dried powders from Examples 1-3 and Comparative Examples 1-10 were prepared into 50 μg / mL solutions with distilled water to construct a DPPH free radical scavenging reaction system, as shown in the table below:

[0176] reaction system a b c Lyophilized powder solution (ml) － 0.5 0.5 0.2mM DPPH methanol solution (ml) 0.5 0.5 － Deionized water (ml) 0.5 － 0.5

[0177] After sample addition, the sample was placed in a constant temperature and dark place at 25℃ for 30 min, and the absorbance values ​​of a, b, and c were measured at 517 nm. Each sample was repeated 5 times. The formula for calculating the DPPH free radical scavenging capacity is as follows:

[0178] DPPH free radical scavenging rate = [1 - (bc) / a] × 100%

[0179] Table 2:

[0180] Grouping DPPH free radical scavenging rate (%) Example 1 96.1±1.2 Example 2 90.1±0.9 Example 3 91.2±2.1 Comparative Example 1 67.1±3.2 Comparative Example 2 68.2±2.1 Comparative Example 3 56.7±3.4 Comparative Example 4 69.6±2.2 Comparative Example 5 66.1±0.9 Comparative Example 6 60.2±2.9 Comparative Example 7 64.2±3.1 Comparative Example 8 62.7±1.3 Comparative Example 9 63.6±1.9 Comparative Example 10 59.7±4.1

[0181] The results in Table 2 show that the DPPH free radical scavenging effects of Examples 1-3 are significantly higher than those of Comparative Examples 1-10. A comparison between Example 1 and Comparative Examples 1-4 indicates that American ginseng and sea buckthorn require co-fermentation with *Saccharomyces cerevisiae* CICC1009 and *Lactobacillus bulgaricus* CICC6047 to exert their antioxidant effects. Comparative Examples 4-5 show that the antioxidant effect of deer antler, without enzymatic hydrolysis by *Bacillus subtilis* protease, is significantly lower than that of Example 1. Comparative Examples 1-10 indicate that fermentation of American ginseng and sea buckthorn with *Saccharomyces cerevisiae* CICC1009 and *Lactobacillus bulgaricus* CICC6047, followed by enzymatic hydrolysis of deer antler with *Bacillus subtilis* protease, and synergistic effects of the fermentation and hydrolysis products with *Rhodiola rosea*, results in a high DPPH free radical scavenging rate.

[0182] 3. Bowel movement test

[0183] Mice: Ninety-six male Kunming mice, weighing 18.0–22.0 g, specific pathogen-free grade, were purchased from Vital Rivers Ltd. The mice were randomly divided into 12 groups: a model control group, a blank control group, and drug-treated groups (Examples 1-3, Comparative Examples 1-4, Comparative Examples 7-8, and Comparative Example 11), with eight mice in each group.

[0184] Specific experimental methods:

[0185] The American ginseng compound freeze-dried powders of Examples 1-3, Comparative Examples 1-4, Comparative Examples 7-8, and Comparative Example 11 were prepared into a 40 mg / mL solution with distilled water and administered to mice at a gavage volume of 0.2 mL / 10 (g·BW) (i.e., 200 μL of the compound solution per 10 g body weight per day). The model control group and the blank control group were administered the corresponding volume of distilled water at a gavage volume of 0.2 mL / 10 (g·BW) once a day for 7 consecutive days.

[0186] Mice in each group were fasted for 16 hours after the last administration of the test sample, but were allowed free access to water. The model control group, Examples 1-3, and Comparative Examples 1-4, 7-8, and 11 were administered compound diphenoxylate 10 mg / (kg·BW) by gavage, while the blank control group received an equal volume of distilled water. 0.5 hours after administration of compound diphenoxylate, mice in the model control group and blank control group were administered ink by gavage, while mice in Examples 1-3, Comparative Examples 1-4, 7-8, and 11 were administered ink containing the corresponding test sample, and timing was started simultaneously. Each animal was housed individually, with normal access to water and food. The time of the first black stool, the number of stool particles excreted within 5 hours, and their weight were observed and recorded for each mouse. See Table 3 for details.

[0187] Table 3

[0188]

[0189] The results in Table 3 show that the laxative effect of Example 1 is significantly higher than that of Examples 2-3, Comparative Examples 1-4, Comparative Examples 7-8, and Comparative Example 11. The results of Comparative Examples 1-4 and 7-8 indicate that American ginseng and sea buckthorn need to be fermented with Saccharomyces cerevisiae CICC1009 and Lactobacillus bulgaricus CICC6047 to exert a mild laxative effect. The results of Comparative Example 11 indicate that peppermint plays an indispensable role in the laxative effect. In conclusion, American ginseng and sea buckthorn need to be fermented with Saccharomyces cerevisiae CICC1009 and Lactobacillus bulgaricus CICC6047. The freeze-dried powder produced after fermentation and the freeze-dried peppermint powder can work synergistically to exert a mild laxative effect.

[0190] The application of the American ginseng compound freeze-dried powder provided in this application in the preparation of products for prolonging life, anti-aging and bowel-regulating.

[0191] The embodiments described above are merely illustrative of specific implementations of the present invention, and while the descriptions are detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention.

Claims

1. A compound freeze-dried powder of American ginseng, characterized in that, The freeze-dried powder is composed of the following components by weight: 25-35 parts of freeze-dried American ginseng powder and 5-15 parts of freeze-dried sea buckthorn powder, which are fermented together; 15-25 parts of Rhodiola rosea powder; 10-20 parts of freeze-dried deer antler powder, which are enzymatically hydrolyzed together; 3-8 parts of Cornus officinalis powder; 2-5 parts of Allium macrostemon powder; 1-3 parts of Mentha haplocalyx powder; and 2-5 parts of Lophatherum gracile powder. The freeze-dried American ginseng powder and freeze-dried sea buckthorn powder are mixed in a certain proportion and then fermented successively with Lactobacillus bulgaricus CICC6047 and Saccharomyces cerevisiae CICC1009. The freeze-dried deer antler powder is enzymatically hydrolyzed with Bacillus subtilis protease.

2. The American ginseng compound freeze-dried powder according to claim 1, characterized in that, The freeze-dried powder is composed of the following components by weight: 30 parts of American ginseng freeze-dried powder and 10 parts of sea buckthorn freeze-dried powder, which are fermented to obtain freeze-dried powder A; 20 parts of Rhodiola rosea; 15 parts of deer antler freeze-dried powder, which are obtained by enzymatic hydrolysis to obtain freeze-dried powder B; 5 parts of Cornus officinalis; 3 parts of Allium macrostemon; 2 parts of Mentha haplocalyx; and 3 parts of Lophatherum gracile.

3. A method for preparing American ginseng compound freeze-dried powder according to any one of claims 1-2, characterized in that, Includes the following steps: (1) Fresh American ginseng and sea buckthorn were respectively made into American ginseng freeze-dried powder and sea buckthorn freeze-dried powder. Then, 25-35 parts of American ginseng freeze-dried powder, 5-15 parts of sea buckthorn freeze-dried powder, 2.5 parts of food-grade glucose, 0.01 parts of magnesium sulfate and 20 parts of distilled water were mixed and inoculated with 5% (v / w) Lactobacillus bulgaricus CICC6047 and allowed to ferment at 37℃ for 24 hours. Then, 3% (v / w) Saccharomyces cerevisiae CICC1009 was inoculated and stirred at 30℃ for 48 hours to produce freeze-dried powder A. The inoculation amount was calculated based on the total mass of American ginseng freeze-dried powder and sea buckthorn freeze-dried powder. (2) Weigh out Bacillus subtilis protease, dissolve it in PBS solution at pH 7.1, add deer antler freeze-dried powder for enzymatic hydrolysis for 3 hours at a temperature of 52°C, and then prepare freeze-dried powder B. (3) The remaining raw materials are made into freeze-dried powders, and then compounded with freeze-dried powder A obtained in step (1) and freeze-dried powder B obtained in step (2) in proportion to obtain American ginseng compound freeze-dried powder.

4. A method for preparing a compound freeze-dried powder of American ginseng according to claim 3, characterized in that, The OD600 of the Lactobacillus bulgaricus CICC6047 bacterial culture inoculated in step (1) was 1.0~1.2; the OD600 of the Saccharomyces cerevisiae CICC1009 bacterial culture inoculated in step (1) was 1.0~1.

2.

5. A method for preparing a compound freeze-dried powder of American ginseng according to claim 3, characterized in that, In the enzymatic hydrolysis system of step (2), the enzyme activity of the Bacillus subtilis protease is 150 U / mg, the amount of enzyme added is 0.05% of the freeze-dried deer antler powder, and the mass fraction of the freeze-dried deer antler powder is 15% (w / v).

6. A compound freeze-dried powder of American ginseng according to any one of claims 1-2, characterized in that, The freeze-dried powder can be taken orally after being brewed.

7. A compound freeze-dried powder of American ginseng according to any one of claims 1-2, characterized in that, The freeze-dried powder has the effects of prolonging the lifespan of nematodes, anti-oxidation, and relieving constipation.

8. The use of the American ginseng compound freeze-dried powder according to any one of claims 1 to 2 in the preparation of anti-aging and laxative drugs.

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