A high-activity ginseng fermented product and its application

Abstract

The present invention discloses a highly active ginseng fermented product and its application. Ginseng is fermented using a composite strain composed of independently screened plant lactobacillus Byf‑r‑01 and grape juice spore Hansen yeast Byf‑j‑01. The composite strain metabolic enzyme system is highly complementary, and the prototype ginsenosides are efficiently converted into rare ginsenosides in a collaborative manner, while simultaneously improving the content of polyphenols, flavonoids, alkaloids and volatile oils. The present invention adds chitosan-polyaspartic acid complex to neutralize lactic acid and maintain pH stability. At the same time, it promotes the proliferation of composite strains as a nitrogen source, significantly improves strain biomass and the activity of saponin-related converting enzymes, and promotes the conversion of ginsenosides. The highly active ginseng fermented product prepared by the present invention has both strong antioxidant and anti-fatigue functions. It has been confirmed by animal experiments that it can prolong the exhaustive swimming time of mice, reduce blood urea nitrogen levels, increase liver / muscle glycogen reserves, and significantly alleviate lactic acid accumulation after exercise, providing a new path for the development of high value-added functional products for ginseng.

Description

Technical Field

[0001] The invention belongs to the technical field of ginseng fermentation, and particularly relates to a high-activity ginseng fermentation product and application thereof. Background Art

[0002] Ginseng, also known as the "king of herbs", is a highly respected precious medicinal material in traditional Chinese medicine. It has a long history and extensive medicinal value. It is rich in active polysaccharides, ginsenosides, alkaloids, volatile oils, etc., and has antioxidant, immunity-enhancing, anti-fatigue, anti-aging, blood sugar and blood pressure regulating effects.

[0003] Ginsenosides are the main active ingredients of ginseng. They are divided into unmodified ginsenosides (such as Rb1, Rd, and Re) and rare ginsenosides (such as Rg3, Rh1, and Rh2). Unmodified ginsenosides account for approximately 80% of the total ginsenoside content, but unmodified ginsenosides have low physiological activity and are difficult to absorb and utilize by the human body. Compared with unmodified ginsenosides, rare ginsenosides have better physiological functions and are more easily absorbed by the body. However, the content of rare ginsenosides in ginseng is very small. Increasing the content of rare ginsenosides and achieving the enrichment and extraction of ginseng's active ingredients have been a research hotspot. Therefore, the transformation of unmodified ginsenosides and the enhancement of other bioactive components in ginseng can not only enhance the therapeutic value of ginseng products but also broaden the application of the ginseng industry. Summary of the Invention

[0004] Technical Problem to be Solved: In response to the above technical problems, the present invention is directed to a highly active ginseng fermented product and its application. Ginseng is fermented using a composite strain of independently screened Lactobacillus plantarum Byf-r-01 and Hansenula vitis spores Byf-j-01. The metabolic enzyme systems of the composite strains are highly complementary, synergistically converting prototype ginsenosides into rare ginsenosides while simultaneously increasing the content of polyphenols, flavonoids, alkaloids, and volatile oils. The present invention adds a chitosan-polyaspartic acid complex to neutralize lactic acid and maintain pH stability, while also serving as a nitrogen source to promote the proliferation of the composite strain, significantly increasing strain biomass and the activity of saponin-related converting enzymes, and promoting the conversion of ginsenosides. The highly active ginseng fermented product prepared by the present invention has both strong antioxidant and anti-fatigue properties. Animal experiments have shown that it can prolong the exhaustive swimming time of mice, reduce blood urea nitrogen levels, increase liver / muscle glycogen reserves, and significantly alleviate post-exercise lactic acid accumulation, providing a new path for the development of high-value-added functional ginseng products.

[0005] Technical solution: A high-activity ginseng fermented product, which is made by fermenting ginseng with a composite strain consisting of Lactobacillus plantarum Byf-r-01 and Hansenula vitis spores Byf-j-01; the Lactobacillus plantarum ( Lactobacillus plantarum) Byf-r-01 was deposited in Guangdong Provincial Microbial Culture Collection Center on January 20, 2025, with the deposit number GDMCC NO: 65834; the grape juice Hansenula sporangiophora ( Hanseniaspora uvarum )Byf-j-01 was deposited in Guangdong Provincial Microbiological Culture Collection on January 20, 2025, with the deposit number GDMCC NO: 65833.

[0006] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0007] S1. Select good-quality ginseng (free of diseased spots or mildew), ultrafinely grind to a particle size of 0.5-1 μm to obtain ginseng powder, add water to form a mixed solution, and sonicate to obtain a ginseng homogenate;

[0008] S2. Lactobacillus plantarum Byf-r-01 was activated to prepare Lactobacillus plantarum Byf-r-01 culture liquid, and Hansenula vitis spores Byf-j-01 was activated to prepare Hansenula vitis spores Byf-j-01 culture liquid;

[0009] S3. A bacterial solution of Lactobacillus plantarum Byf-r-01 and a bacterial solution of Hansenula vitis spores Byf-j-01 were mixed to prepare a composite bacterial solution, the composite bacterial solution and a chitosan-polyaspartic acid complex were added to a ginseng homogenate and fermented, filtered and sterilized, and freeze-dried to obtain a highly active ginseng fermentation product.

[0010] Furthermore, the ratio of ginseng powder to water in S1 is 1 g: (4.5-8) mL; and the ultrasonic treatment conditions are ultrasonic power 200-600 W and ultrasonic time 10-20 min.

[0011] Furthermore, the preparation method of the Lactobacillus plantarum Byf-r-01 bacterial liquid in S2 is as follows: Lactobacillus plantarum Byf-r-01 is inoculated into MRS liquid culture medium at an inoculum size of 2-5%, and statically cultured at 30-37°C for 12-15 hours to obtain Lactobacillus plantarum Byf-r-01 seed liquid; 1-5% Lactobacillus plantarum Byf-r-01 seed liquid is inoculated into MRS liquid culture medium, expanded cultured at 30-37°C and 150-200 rpm for 20-24 hours, centrifuged and resuspended with sterile water to obtain Lactobacillus plantarum Byf-r-01 bacterial liquid.

[0012] Furthermore, the preparation method of the Hansenula sporangioides Byf-j-01 bacterial liquid in S2 is as follows: inoculating Hansenula sporangioides Byf-j-01 into YPD liquid culture medium at an inoculum size of 1.5-5%, culturing at 28-32° C. for 12-15 hours to obtain a Hansenula sporangioides Byf-j-01 seed liquid; inoculating 1-5% of the Hansenula sporangioides Byf-j-01 seed liquid into YPD liquid culture medium, expanding the culture at 28-32° C. and 150-200 rpm for 20-24 hours, centrifuging, and resuspending with sterile water to obtain a Hansenula sporangioides Byf-j-01 bacterial liquid.

[0013] Furthermore, the volume ratio of the Lactobacillus plantarum Byf-r-01 bacterial solution and the Hansenula vitis spore Byf-j-01 bacterial solution in the S3 is (1.5-5): (1-3.5); the addition amount of the composite bacterial solution is 2.5-6vt%; the bacterial concentration of the composite bacterial strain is (1.5-7)×10 8 CFU / mL.

[0014] Furthermore, the preparation method of the chitosan-polyaspartic acid complex in S3 is as follows: chitosan and polyaspartic acid are uniformly mixed in a mass ratio of (1-4):1 and dissolved in water, treated at 20-30°C for 1-3h, and dried to obtain the chitosan-polyaspartic acid complex.

[0015] Furthermore, the added amount of chitosan-polyaspartic acid complex in S3 is 2.5-5 wt%.

[0016] Furthermore, the specific conditions for fermentation in S3 are a fermentation temperature of 30-35°C and a fermentation time of 1-4 days; the specific operation of the freeze-drying is: placing the filtered and sterilized ginseng fermented liquid in a vacuum freeze dryer for freeze drying, and the specific conditions for freeze drying are a pre-freezing temperature of -40--30°C, a pre-freezing time of 1-3h, a heating temperature of 10-15°C, a drying time of 12-24h, and a vacuum degree of 10-20kPa.

[0017] The application of the above-mentioned high-activity ginseng fermented product in the preparation of anti-fatigue drugs.

[0018] Furthermore, the forms of the medicine include tablets, capsules, granules, and granules.

[0019] Beneficial effects

[0020] 1. The present invention uses a composite bacterial strain composed of self-screened strains (Lactobacillus plantarum Byf-r-01 and Hansenula vitis sporangium Byf-j-01) to ferment ginseng. The biological enzyme system produced by their metabolism has highly complementary properties and can convert the prototype ginsenosides contained in ginseng into rare ginsenosides, thereby increasing the rare ginsenoside content of highly active ginseng fermented products; at the same time, it also increases the content of polyphenols, flavonoids, and alkaloids. In addition, the cellulase produced by the bacterial strain metabolism enzymatically degrades the cellulose in the ginseng cell wall, releasing bound volatile oil molecules, thereby increasing the volatile oil content, and further significantly improving the active ingredients of the ginseng fermented products, resulting in highly active ginseng fermented products.

[0021] 2. During the preparation of the highly active ginseng fermented product of the present invention, a chitosan-polyaspartic acid complex is added. The chitosan-polyaspartic acid complex is prepared by forming an amide bond between the amino group of chitosan and the carboxyl group of polyaspartic acid. The chitosan-polyaspartic acid complex can neutralize the lactic acid produced by the metabolism of Lactobacillus plantarum Byf-r-01, maintaining the pH stability of the fermentation system. In addition, the chitosan-polyaspartic acid complex can be absorbed and utilized as a nitrogen source by Lactobacillus plantarum Byf-r-01 and Hansenula vitis spores Byf-j-01, providing nutrients and energy for the fermentation metabolism of the strains, promoting the proliferation of Lactobacillus plantarum Byf-r-01 and Hansenula vitis spores Byf-j-01, significantly increasing their biomass and activity, and indirectly increasing the synthesis amount and activity of enzyme systems related to the conversion of rare ginsenosides, ultimately efficiently converting them into a variety of rare ginsenosides.

[0022] 3. The high-activity ginseng fermented product prepared by the present invention has high antioxidant capacity and Fe 3+ It has the reducing ability and can significantly scavenge DPPH free radicals, ABTS+ free radicals, hydroxyl free radicals and superoxide anion free radicals; on the other hand, it has good anti-fatigue function, which can significantly prolong the exhaustive swimming time of mice, significantly reduce the blood urea nitrogen level of mice after weight-bearing swimming, increase the reserves of liver glycogen and muscle glycogen in the body, increase the content of lactate dehydrogenase in the body of mice, and reduce the accumulation of lactic acid after exercise.

[0023] 4. The present invention uses Lactobacillus plantarum Byf-r-01 and Hansenula vitis spores Byf-j-01 for composite fermentation to prepare a highly active ginseng fermented product, which has both high activity and good antioxidant and anti-fatigue functions. It can provide a theoretical basis and data support for the development and application of ginseng functional products. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The rare ginsenoside content in Example 2, Comparative Examples 1-4 and the sample before fermentation;

[0025] Figure 2 is the free radical scavenging rate of Example 2 and Comparative Examples 1-4, wherein the free radicals include DPPH•, ABTS+•, •OH, O2- •;

[0026] Figure 3 The exhausted swimming time, urea nitrogen content, liver glycogen content and muscle glycogen content of mice in Example 2 and Comparative Examples 1-4 groups are shown, where A represents the exhausted swimming time, B represents the urea nitrogen content, C represents the liver glycogen content, and D represents the muscle glycogen content;

[0027] Figure 4 The lactate dehydrogenase content of mice in Example 2 and Comparative Examples 1-4 groups is shown. DETAILED DESCRIPTION

[0028] The present invention will be further described below in conjunction with examples, which are provided to explain the present invention and are not limited to the following examples:

[0029] The strains used in the present invention are: 1. Lactobacillus plantarum ( Lactobacillus plantarum ) Byf-r-01: Deposit number GDMCC NO: 65834, deposit date January 20, 2025, deposited by Guangdong Provincial Microbial Culture Collection Center; ② Hansenula sporangiophora vitis vinifera ( Hanseniaspora uvarum )Byf-j-01: Deposition number GDMCC NO: 65833, deposition date January 20, 2025, deposition unit Guangdong Provincial Microbiological Culture Collection Center.

[0030] Example 1

[0031] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0032] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0033] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0034] ① Inoculate 5 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, and incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0035] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0036] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0037] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.50 × 10 8 CFU / mL), 12 mL of the composite bacterial solution and 9 g of chitosan-polyaspartic acid complex were added to 300 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0038] Example 2

[0039] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0040] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0041] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0042] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0043] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0044] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0045] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.55 × 10 8 CFU / mL), 10 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0046] Example 3

[0047] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0048] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0049] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0050] ① Inoculate 5 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, and incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0051] ② Inoculate 10 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 7.5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0052] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0053] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.50 × 10 8 CFU / mL), 12 mL of the composite bacterial solution and 9 g of chitosan-polyaspartic acid complex were added to 300 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0054] Example 4

[0055] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0056] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0057] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0058] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0059] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0060] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0061] S4. Mix 15 mL of Lactobacillus plantarum Byf-r-01 and 35 mL of Hansenula vitis spores Byf-j-01 to prepare a composite bacterial solution (with a bacterial concentration of 4.65 × 10 8 CFU / mL), 10 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0062] Example 5

[0063] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0064] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0065] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0066] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0067] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0068] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0069] S4. Mix 40 mL of Lactobacillus plantarum Byf-r-01 and 10 mL of Hansenula vitis spores Byf-j-01 to prepare a composite bacterial solution (with a bacterial concentration of 4.55 × 10 8 CFU / mL), 12 mL of the composite bacterial solution and 9 g of chitosan-polyaspartic acid complex were added to 300 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0070] Example 6

[0071] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0072] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0073] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0074] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0075] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0076] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0077] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.50 × 10 8 CFU / mL), 10 mL of the composite bacterial solution and 12 g of chitosan-polyaspartic acid complex were added to 400 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0078] Example 7

[0079] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0080] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0081] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0082] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0083] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0084] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0085] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.51 × 10 8 CFU / mL), 15 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0086] Example 8

[0087] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0088] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0089] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0090] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0091] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0092] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0093] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.50 × 10 8 CFU / mL), 12 mL of the composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 300 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0094] Example 9

[0095] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0096] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0097] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0098] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0099] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0100] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0101] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.53 × 10 8 CFU / mL), 10 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 35°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0102] Example 10

[0103] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0104] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0105] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0106] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0107] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0108] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0109] S4. Mix 30 mL of Lactobacillus plantarum Byf-r-01 culture and 20 mL of Hansenula vitis spores Byf-j-01 culture to prepare a composite culture (with a strain concentration of 4.50 × 10 8 CFU / mL), 10 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 32°C for 1.5 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0110] Comparative Example 1

[0111] The difference between this comparative example and Example 2 is that the chitosan-polyaspartic acid complex is not added.

[0112] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0113] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0114] S2. Activation of Lactobacillus plantarum Byf-r-01 and Hansenula sporogenes Byf-j-01

[0115] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0116] ② Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0117] S3. Mix 30 mL of Lactobacillus plantarum Byf-r-01 and 20 mL of Hansenula vitis spores Byf-j-01 to prepare a composite bacterial solution (with a bacterial concentration of 4.55 × 10 8 CFU / mL), 10 mL of the composite bacterial solution was added to 250 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0118] Comparative Example 2

[0119] The difference between this comparative example and Example 2 is that only Hansenula sporangiophora vitis vinifera Byf-j-01 was used.

[0120] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0121] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0122] S2. Activation of Hansenula sporangiophora Byf-j-01

[0123] ① Inoculate 5 mL of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, and culture at 30°C for 15 h to obtain a seed solution of Hansenula sporangiophora Byf-j-01; inoculate 5 mL of the seed solution of Hansenula sporangiophora Byf-j-01 into 250 mL of YPD liquid medium, expand the culture at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporangiophora Byf-j-01;

[0124] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0125] S4. Add 10 mL of Hansenula vitis spores Byf-j-01 bacterial solution (with a bacterial concentration of 4.50 × 10 8 CFU / mL) and 7.5 g of chitosan-polyaspartic acid complex were fermented at 32 °C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35 °C, pre-freezing time 2.5 h, heating temperature 15 °C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0126] Comparative Example 3

[0127] The difference between this comparative example and Example 2 is that only Lactobacillus plantarum Byf-r-01 is used.

[0128] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0129] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0130] S2. Activation of Lactobacillus plantarum Byf-r-01

[0131] ① Inoculate 10 mL of Lactobacillus plantarum Byf-r-01 into 250 mL of MRS liquid culture medium, incubate at 37°C for 12 h to obtain a Lactobacillus plantarum Byf-r-01 seed solution; inoculate 7.5 mL of Lactobacillus plantarum Byf-r-01 seed solution into 250 mL of MRS liquid culture medium, incubate at 37°C and 200 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a Lactobacillus plantarum Byf-r-01 bacterial solution;

[0132] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0133] S4. Add 10 mL of Lactobacillus plantarum Byf-r-01 bacterial solution (the bacterial concentration is 4.50 × 10 8 CFU / mL) and 7.5 g of chitosan-polyaspartic acid complex were fermented at 32 °C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35 °C, pre-freezing time 2.5 h, heating temperature 15 °C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0134] Comparative Example 4

[0135] The difference between this comparative example and Example 2 is that commercially available Lactobacillus plantarum and Hansenula vitis spores were used, wherein Lactobacillus plantarum M32-2 (CICC20314) and Hansenula vitis spores F30 (CICC 31872) were both purchased from the China Industrial Microbiological Culture Collection Center.

[0136] A method for preparing a high-activity ginseng fermented product comprises the following steps:

[0137] S1. Select good quality ginseng, ultrafinely grind to obtain ginseng powder, add 500mL of water to 100g of ginseng powder to make a mixed solution, and ultrasonically treat at an ultrasonic power of 450W for 15min to obtain a ginseng homogenate;

[0138] S2. Activation of Lactobacillus plantarum M32-2 and Hansenula sporogenes F30

[0139] ① Inoculate 10 mL of Lactobacillus plantarum M32-2 into 250 mL of MRS liquid culture medium, incubate at 37° C. for 12 h to obtain a Lactobacillus plantarum M32-2 seed solution; inoculate 7.5 mL of the Lactobacillus plantarum M32-2 seed solution into 250 mL of MRS liquid culture medium, incubate at 37° C. and 200 rpm for 24 h, centrifuge, and resuspend with sterile water to obtain a Lactobacillus plantarum M32-2 bacterial solution;

[0140] ② Inoculate 5 mL of Hansenula sporogenes F30 into 250 mL of YPD liquid medium, incubate at 30°C for 15 h to obtain a seed solution of Hansenula sporogenes F30; inoculate 5 mL of the seed solution of Hansenula sporogenes F30 into 250 mL of YPD liquid medium, incubate at 30°C and 150 rpm for 24 h, centrifuge, and resuspend in sterile water to obtain a culture solution of Hansenula sporogenes F30;

[0141] S3. Weigh 30 g of chitosan and 10 g of polyaspartic acid, mix well and dissolve in 300 mL of water, treat at 25 ° C for 1.5 h, and dry to obtain a chitosan-polyaspartic acid complex;

[0142] S4. Mix 30 mL of Lactobacillus plantarum M32-2 culture and 20 mL of Hansenula viticola F30 culture to prepare a composite culture (with a strain concentration of 4.55 × 10 8 CFU / mL), 10 mL of composite bacterial solution and 7.5 g of chitosan-polyaspartic acid complex were added to 250 mL of ginseng homogenate, fermented at 32°C for 3 days, filtered and sterilized, and freeze-dried in a vacuum freeze dryer (pre-freezing temperature -35°C, pre-freezing time 2.5 h, heating temperature 15°C, drying time 24 h, vacuum degree 18 kPa) to obtain a highly active ginseng fermented product.

[0143] Performance Testing

[0144] (1) Polysaccharide content

[0145] Glucose was used as a standard substance and the polysaccharide content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined by the phenol-sulfuric acid method.

[0146] (2) Total phenol content

[0147] Gallic acid was used as a standard substance and the total phenol content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined by the Folin-phenol colorimetric method.

[0148] (3) Total flavonoid content

[0149] The total flavonoids content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined by aluminum chloride colorimetry using rutin as a standard.

[0150] (4) Total saponin content

[0151] Using ginsenoside Re as a standard, the total saponin content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined by the vanillin-glacial acetic acid method.

[0152] (5) Alkaloid content

[0153] The total alkaloid content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined by acid dye colorimetry.

[0154] (6) Volatile oil content

[0155] The volatile oil content in the highly active ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4 was determined according to the volatile oil determination method in Part 4, 2204 of the Chinese Pharmacopoeia 2020 Edition.

[0156] Table 1 Physical and chemical indicators of the high-activity ginseng fermented products prepared in Examples 1-10 and Comparative Examples 1-4

[0157]

[0158] As shown in Table 1, the polysaccharide and total saponin contents of Examples 1-10 and Comparative Examples 1-4 were lower than those of the pre-fermentation samples. This is because the growth and metabolism of Lactobacillus plantarum and Hansenula vitis spores require polysaccharides in ginseng as a carbon source, and the abundant enzymes produced by metabolism convert the prototype ginsenosides into rare ginsenosides, so the polysaccharide and total saponin contents decreased. The total phenols, total flavonoids, alkaloids and volatile oils of Examples 1-10 were higher than those of the pre-fermentation samples, and were also higher than those of Comparative Examples 1-4, indicating that the Lactobacillus plantarum Byf-r used in the examples -1 and Hansenula sporogenes Byf-j-1 can metabolize the macromolecular substances in ginseng and efficiently convert them into polyphenols and flavonoids. At the same time, the chitosan-polyaspartic acid complex can not only serve as a nitrogen source to provide energy for the bacteria, but also neutralize the lactic acid produced by Lactobacillus plantarum Byf-r-1 to maintain the pH stability of the fermentation system, increase the activity of related enzymes, and promote the production of alkaloids and volatile oils in highly active ginseng fermented products, thereby increasing the bioactive substances in highly active ginseng fermented products and giving them excellent biological benefits.

[0159] (7) Quantitative analysis of rare ginsenosides

[0160] The rare ginsenosides in the highly active ginseng fermented products prepared in Example 2 and Comparative Examples 1-4 were analyzed according to GB / T 41726-2022 Identification and Detection Method of Ginseng Monomer Saponins.

[0161] Depend on Figure 1 It can be seen that the rare ginsenosides Rd, Rg3, Rh1, Rh2 and Rh4 in the highly active ginseng fermented products prepared in Example 2 and Comparative Examples 1-4 are all at a high level, and the rare ginsenosides in Example 2 are all higher than those in Comparative Examples 1-4, indicating that on the one hand, the combined use of the self-screened bacteria (Lactobacillus plantarum Byf-r-1 and Hansenula vitis spores Byf-j-1) has a highly complementary biological enzyme system produced by its metabolism, which efficiently decomposes the prototype ginsenosides in ginseng into rare ginsenosides; on the other hand, the addition of the chitosan-polyaspartic acid complex can not only provide the nutrients and energy required by the composite bacteria as a nitrogen source, but also can improve the activity of the enzyme system related to the conversion of the prototype ginsenosides into rare ginsenosides, thereby increasing the production of rare ginsenosides.

[0162] Example 11

[0163] Verification test of antioxidant capacity of high-activity ginseng fermentation products

[0164] The DPPH free radical scavenging ability, ABTS cation free radical scavenging ability, hydroxyl free radical scavenging ability, superoxide anion free radical scavenging ability and Fe free radical scavenging ability of the high-activity ginseng fermented products prepared in Example 2 and Comparative Examples 1-4 were respectively measured. 3+ The restoring power is as follows:

[0165] (1) DPPH free radical scavenging ability: Prepare a sample solution with a mass concentration of 500 μg / mL and a positive control V C Solution, add 20 μL of sample and positive control solution and 180 μL of DPPH working solution to a 96-well plate, place it in a 37°C thermostat in the dark for 10 minutes, measure the absorbance at 517 nm, the blank group is anhydrous ethanol + DPPH working solution, the background group is sample / positive control Vc solution + anhydrous ethanol, and calculate the DPPH free radical scavenging rate according to the formula; the formula is: DPPH free radical scavenging rate (%) = [1-(A1-A2) / A0] × 100%, where A1 is the absorbance of the sample / positive control group, A2 is the absorbance of the background group of the positive control / sample, and A0 is the absorbance of the blank group;

[0166] (2) ABTS cationic free radical scavenging ability: Prepare 500 μg / mL sample solution and positive control Vc solution, add 20 μL of sample and positive control solution and 180 μL of ABTS working solution into a 96-well plate, set up three parallels for each concentration, place in a 37°C constant temperature box in the dark for 10 minutes, and measure the absorbance of each group at 734 nm; the blank group is distilled water + ABTS working solution, and the background group is sample / positive control Vc solution + distilled water; calculate the ABTS+ free radical scavenging rate according to the formula; the formula is: ABTS+ free radical scavenging rate (%) = [1-(A1-A2) / A0] × 100%, where A1 is the absorbance of the sample / positive control group, A2 is the absorbance of the background group of the positive control / sample, and A0 is the absorbance of the blank group;

[0167] (3) Hydroxyl radical scavenging ability: 1 mL of 1.8 mmol / LFeSO4, 1.8 mmol / L salicylic acid-ethanol solution and sample solution were mixed, and then 1 mL of 1.76 mmol / L H2O2 was added, and the mixture was reacted in a constant temperature water bath at 37°C for 30 min. The absorbance value A1 was measured at 510 nm using an enzyme marker; 1 mL of sample solution and 3 mL of ethanol were mixed, and the absorbance value A0 was measured at 510 nm using an enzyme marker; 1 mL of 1.8 mmol / LFeSO4, 1.8 mmol / L salicylic acid-ethanol solution and ethanol were mixed, and then 1 mL of 1.76 mmol / L H2O2 was added, and the mixture was reacted in a constant temperature water bath at 37°C for 30 min. The absorbance value A2 of the solution system without antioxidant was measured at 510 nm using an enzyme marker, and the hydroxyl radical scavenging rate was calculated according to the formula; the formula is: Hydroxyl radical scavenging rate (%) = [1-(A1-A0) / A2] × 100%;

[0168] (4) Superoxide anion radical scavenging ability: Prepare 1000 μg / mL sample solution and positive control Vc solution, add 20 μL sample solution and 90 μL buffer, 60 μL distilled water, and 10 μL pyrogallic acid solution into a 96-well plate, set up three parallels for each concentration, place in a 25°C constant temperature box in the dark for 20 minutes, and measure the absorbance of each group at 320 nm; the blank group uses 20 μL distilled water and subsequent reaction working solution, and the background group is sample / positive control Vc solution + distilled water. The superoxide anion radical scavenging rate is calculated according to the formula; the formula is: superoxide anion radical scavenging rate (%) = [1-(A1-A2) / A0] × 100%, where A1 is the absorbance of the sample / positive control group, A2 is the absorbance of the background group of the positive control / sample, and A0 is the absorbance of the blank group;

[0169] (5) Total reducing capacity: Prepare 500 μg / mL sample solution and positive control Vc solution, take 300 μL sample into a 2 mL centrifuge tube, then add 300 μL phosphate buffer and 300 μL potassium ferricyanide solution in sequence, shake gently to mix, place in a 50°C constant temperature box to react for 20 minutes, then immediately put into an ice water bath to cool, and then add 300 μL trichloroacetic acid solution to terminate the reaction. Subsequently, each group was centrifuged at 2500 rpm for 5 minutes, and 90 μL of supernatant was taken into a 96-well plate, and 90 μL distilled water and 18 μL ferric chloride solution were added in sequence. The plate was allowed to stand for 10 minutes, and the absorbance of each group was measured at 700 nm; the blank control group used 300 μL distilled water plus subsequent reaction solution, and the positive control used ascorbic acid (Vc); the total reducing capacity was calculated according to the formula: total reducing capacity = A1-A0, where A1 is the absorbance of the sample / positive control group, and A0 is the absorbance of the blank group.

[0170] Table 2 Total reducing capacity of positive control, Example 2 and Comparative Examples 1-4

[0171]

[0172] Depend on Figure 2 As shown in Table 2, the high-activity ginseng fermented product prepared in Example 2 has a strong effect on a variety of free radicals (including DPPH•, ABTS+•, •OH, O2 - •) have good scavenging ability, which is better than that of Comparative Examples 1-4, and the total reducing power is also higher than that of Comparative Examples 1-4. This is because compared with Comparative Examples 1-4, the high-activity ginseng fermented product prepared in Example 2 has more polyphenols and flavonoids, as well as highly active rare ginsenosides, so its antioxidant function is better than that of Comparative Examples 1-4.

[0173] Example 12

[0174] Verification test of the anti-fatigue function of high-activity ginseng fermented products

[0175] The anti-fatigue effect of the highly active ginseng fermented products prepared in Example 2 and Comparative Examples 1-4 was analyzed by animal experiments. Specifically, 70 SPF-grade Kunming mice (male, weighing 18-22 g) were randomly divided into 7 groups (blank control group, positive group, Example 2 group, Comparative Example 1 group, Comparative Example 2 group, Comparative Example 3 group, Comparative Example 4 group), with 10 mice in each group, and adaptive feeding was performed for 3 days; administration method: ① The blank control group was given an equal volume of distilled water, and the administration was continued by gavage for 30 days; ② The positive group was given American ginseng lozenges at a dose of 0.48 g / kg / d; ③ Example 2 group, Comparative Example 1 group, Comparative Example 2 group, Comparative Example 3 group, and Comparative Example 4 group were all given freeze-dried powder of the highly active ginseng fermented product at a dose of 1.95 g / kg / d; feeding method: all groups of mice were placed in the standard experimental animal room for feeding, with an ambient temperature of 25±1℃ and a relative humidity of 50-60%, alternating day and night, and given sufficient sterilized mouse feed and sterilized water. Feed and water were available freely during the feeding period, and the feeding cycle was 30 days; after the end of the feeding cycle, the body weight of each group of mice was weighed, and the mice were subjected to a weighted swimming test to record the exhaustive swimming time of the mice. The mice were then killed and samples were taken to analyze the content of urea nitrogen, liver glycogen, muscle glycogen and lactate dehydrogenase.

[0176] Table 3 Initial and final body weights of mice in each treatment group

[0177]

[0178] As shown in Table 3, compared with the blank control group and the positive group, the high-activity ginseng fermented products prepared in Example 2 and Comparative Examples 1-4 had little effect on the weight gain of mice when fed.

[0179] Fatigue is mainly manifested in the inability to concentrate and decreased exercise endurance. The duration of exhaustion exercise can be used as an effective means to evaluate the strength of exercise ability. Figure 3 As shown in A, compared with the positive group, the exhaustive swimming time of the mice in Example 2 and Comparative Examples 1-3 groups was longer, while the exhaustive swimming time of the mice in Comparative Example 4 group was shorter; Figure 3 B. Figure 3 C and Figure 3 As shown in D, the urea nitrogen content of the mice in Example 2 group was lower than that in the positive group and the comparative example 1-4 groups, while the liver glycogen content and muscle glycogen content were higher than those in the positive group and the comparative example 1-4 groups; Figure 4 It can be seen that the lactate dehydrogenase content of Example 2 group is higher than that of the positive group and the comparative examples 1-4 groups. Therefore, it can be seen that the high-activity ginseng fermented product prepared in Example 2 group has good anti-fatigue ability, can promote the body's glycogen reserves, and reduce the level of urea nitrogen produced after protein metabolism in the body and the accumulation of lactic acid in the body.

[0180] The above description is merely a preferred embodiment of the present invention and does not constitute any form of limitation to the present invention. Any person skilled in the art can, without departing from the spirit and technical solution of the present invention, use the methods and technical contents disclosed above to make many possible changes and modifications to the technical solution of the present invention, or modify it into an equivalent embodiment with equivalent changes. Therefore, any simple modification, equivalent replacement, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention shall still fall within the scope of protection of the technical solution of the present invention.

Claims

1. A high-activity ginseng fermented product, characterized in that: The highly active ginseng fermented product is prepared by fermenting ginseng using a composite strain consisting of Lactobacillus plantarum Byf-r-01 and Hansenula sporangiophora vitis Byf-j-01; the Lactobacillus plantarum Byf-r-01 was deposited in the Guangdong Provincial Center for Microbiological Culture Collection on January 20, 2025, with a deposit number of GDMCC NO: 65834; the Hansenula sporangiophora vitis Byf-j-01 was deposited in the Guangdong Provincial Center for Microbiological Culture Collection on January 20, 2025, with a deposit number of GDMCC NO: 65833; The method for preparing a high-activity ginseng fermented product comprises the following steps: S1. Select good quality ginseng, ultrafinely grind to a particle size of 0.5-1 μm to obtain ginseng powder, add water to mix to form a mixed solution, and ultrasonically treat to obtain a ginseng homogenate; S2. Lactobacillus plantarum Byf-r-01 was activated to prepare Lactobacillus plantarum Byf-r-01 culture liquid, and Hansenula vitis spores Byf-j-01 was activated to prepare Hansenula vitis spores Byf-j-01 culture liquid; S3. A Lactobacillus plantarum Byf-r-01 bacterial solution and a Hansenula vitis spore Byf-j-01 bacterial solution are mixed to form a composite bacterial solution, the composite bacterial solution and a chitosan-polyaspartic acid complex are added to a ginseng homogenate and fermented, filtered and sterilized, and freeze-dried to obtain a highly active ginseng fermented product; the volume ratio of the Lactobacillus plantarum Byf-r-01 bacterial solution to the Hansenula vitis spore Byf-j-01 bacterial solution is (1.5-5):(1-3.5); the amount of the composite bacterial solution added is 2.5-6% by volume; and the bacterial concentration of the composite strain is (1.5-7)×10 8 CFU / mL.

2. The high-activity ginseng fermented product according to claim 1, characterized in that: The ratio of ginseng powder to water in S1 is 1 g: (4.5-8) mL; the ultrasonic treatment conditions are ultrasonic power 200-600 W and ultrasonic time 10-20 min.

3. The high-activity ginseng fermented product according to claim 1, characterized in that: The preparation method of the chitosan-polyaspartic acid complex in S3 is as follows: chitosan and polyaspartic acid are uniformly mixed in a mass ratio of (1-4):1 and dissolved in water, treated at 20-30°C for 1-3h, and dried to obtain the chitosan-polyaspartic acid complex.

4. The high-activity ginseng fermented product according to claim 1, characterized in that: The added amount of the chitosan-polyaspartic acid complex in S3 is 2.5-5 wt %.

5. The high-activity ginseng fermented product according to claim 1, characterized in that: The specific conditions of the fermentation in S3 are a fermentation temperature of 30-35° C. and a fermentation time of 1-4 days.

6. Use of the highly active ginseng fermented product according to claim 1 in the preparation of anti-fatigue drugs.

7. The use according to claim 6, characterized in that The forms of the medicine include tablets, capsules, granules and granules.

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