A composition for preventing and treating high altitude oxidative stress spermatogenic dysfunction and a preparation method thereof

Abstract

The application relates to the technical field of biological medicine, and particularly discloses a composition for resisting highland oxidative stress spermatogenic disorder and a preparation method thereof. The composition for resisting highland oxidative stress spermatogenic disorder comprises the following raw materials in parts by weight: wolfberry, rhizoma polygonati, semen cuscutae, angelica sinensis, astragalus, raspberry, Akkermansia muciniphila bacterial powder, a TRPV4 inhibitor and an excipient. The preparation method comprises the following steps: S1, pretreatment of raw materials; S2, preparation of traditional Chinese medicine extracts; S3, mixing and blending; and S4, preparation of preparations. Through the technical scheme of ternary synergistic matching of a traditional Chinese medicine compound, functional probiotics and a targeted inhibitor, the technical effects of precise intervention on the whole chain of highland oxidative stress spermatogenic disorder, realization of three-in-one comprehensive protection of antioxidation, regulation of flora and prevention of immune damage are achieved, and the problems of single target effect, inability to cover multi-link damage of the intestine and testis axis, limited curative effect and easy recurrence are solved.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to a composition for combating spermatogenesis disorders caused by high-altitude oxidative stress and its preparation method. Background Technology

[0002] The high-altitude environment, characterized by low pressure and low oxygen, easily triggers oxidative stress in the body. This stress remotely regulates spermatogenesis through the gut-testis axis, creating a vicious cycle of gut microbiota imbalance, abnormal accumulation of metabolic products, and testicular immune damage, ultimately leading to spermatogenic dysfunction and affecting the reproductive health of people exposed to high altitudes. Existing intervention products to address this issue mainly include single antioxidants, hormone supplements, and traditional Chinese medicine formulas. Single antioxidants focus on scavenging oxygen free radicals, hormone supplements emphasize direct regulation of reproductive endocrine function, and traditional Chinese medicine formulas are mostly designed to tonify the kidneys and promote spermatogenesis or provide mild antioxidant effects—all interventions targeting single links in the pathological chain.

[0003] Current technologies lack a full understanding of the gut-testis axis-mediated effects and have not developed a comprehensive intervention system covering multiple aspects such as oxidative stress, gut microbiota imbalance, and immune damage. Single-target intervention programs can only temporarily alleviate local symptoms and cannot break the vicious cycle of the pathological chain, resulting in limited efficacy and easy relapse. They are unable to fundamentally solve the spermatogenesis disorders caused by high-altitude oxidative stress, and there is an urgent need to develop highly effective intervention products with multi-target and full-chain synergistic effects. Summary of the Invention

[0004] To address the shortcomings of existing technologies, this invention provides a composition for combating spermatogenesis disorders caused by high-altitude oxidative stress and its preparation method. This solves the problems of existing technologies using single antioxidants, hormone supplements, or traditional Chinese medicine compound formulas, which result in limited efficacy, inability to cover multiple links of damage to the intestinal and testicular axis, and easy relapse.

[0005] To achieve the above objectives, the present invention is implemented through the following technical solution: a composition for resisting high-altitude oxidative stress and spermatogenesis disorders, comprising the following raw materials in parts by weight: 15-25 parts of wolfberry, 12-20 parts of polygonatum, 10-18 parts of dodder seed, 8-16 parts of angelica, 10-20 parts of astragalus, 10-18 parts of raspberry, 2-5 parts of Akkermansia muciniphila mycelium powder, and 0.01-0.05 parts of TRPV4 inhibitor.

[0006] Preferably, the composition further includes excipients selected from at least one of maltodextrin and magnesium stearate, wherein the weight of the excipients accounts for 10-20% of the total weight of the composition.

[0007] Preferably, the TRPV4 inhibitor is HC-067047; the Akkermansiamuciniphila mycelium powder is obtained through an aseptic preparation process; the wolfberry, polygonatum, dodder seed, angelica, astragalus, and raspberry are all dried, mold-free, and impurity-free raw materials, wherein the wolfberry is the dried fruit, the polygonatum is the dried product, the dodder seed is the dried seed, the angelica is the dried root, the astragalus is the dried root, and the raspberry is the dried fruit.

[0008] A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes includes the following steps: S1. Raw material pretreatment: The Chinese herbal raw materials are removed of impurities, washed, dried and then pulverized to obtain mixed Chinese herbal powder; S2. Preparation of Chinese herbal extract: Add extraction solvent to the mixed Chinese herbal powder and soak it for multiple extractions. Combine the extracts and filter them. Concentrate the filtered extract under reduced pressure to obtain an extract paste. Then, vacuum dry the extract paste and pulverize it to obtain Chinese herbal extract powder. S3. Mixing and blending: Under sterile conditions, first premix the Chinese herbal extract powder evenly, then add Akkermansiamuciniphila bacterial powder under preset temperature and humidity conditions, followed by TRPV4 inhibitor and excipients, and mix until the preset homogeneity standard is met. S4. Formulation: The mixture is processed into granules, capsules or tablets using the corresponding process.

[0009] Preferably, in step S1, the drying process employs a hot air circulation drying method, with a drying temperature of 45-50℃ and a drying time of 3-5 hours, resulting in a moisture content of ≤8% for the dried Chinese herbal raw materials. The powder is pulverized using a universal pulverizer. The resulting powder has a particle size of 80-100 mesh. After pulverization, the powder is screened through a vibrating screen to remove coarse powder that does not meet the particle size requirements before mixing to obtain a uniformly mixed powder.

[0010] Preferably, in step S2, the extraction solvent is deionized water, the amount of extraction solvent added is 10-15 times the volume of the mixed Chinese herbal powder, the soaking time is 2-3 hours, and the mixture is stirred once every 30 minutes for 1 minute each time during the soaking process; The extraction was performed twice using a gentle boiling process, with the first extraction lasting 2 hours and the second extraction lasting 1.5 hours. The vacuum concentration was achieved using a rotary evaporator under the following conditions: pressure -0.08 to -0.09 MPa and temperature 55-65°C. The resulting extract had a relative density of 1.15-1.20 at 60°C. Vacuum drying is performed using a vacuum drying oven under the following conditions: temperature 50-55℃, pressure -0.07~-0.08MPa. The moisture content of the dried extract is ≤5%. The dried extract is then pulverized using an ultra-micro pulverizer to obtain a traditional Chinese medicine extract powder with a particle size of 100-120 mesh.

[0011] Preferably, in step S3, the sterile environment is constructed by ultraviolet disinfection for more than 30 minutes, with a cleanliness level of Class 100; the preset temperature and humidity conditions are temperature ≤25℃ and humidity ≤45%. The premixing process uses a three-dimensional motion mixer with a mixing speed of 200-300 r / min and a premixing time of 15-20 minutes; After adding Akkermansiamuciniphila powder, premix manually for 5 minutes, then add TRPV4 inhibitor and excipients, and transfer to a double helix cone mixer for mixing at a speed of 150-250 r / min for 25-35 minutes. The preset homogeneity standard is that the relative standard deviation (RSD) of the mixture is ≤3%; In step S4, the capsules are filled using a fully automatic capsule filling machine with a specification of 0.3-0.5g / capsule. During the filling process, the filling volume difference is sampled and tested every 10 minutes, and the filling volume difference limit is ±5%.

[0012] Preferably, in step S2, the extract filtration adopts a two-stage filtration method: first, large particles of medicinal residue are removed by passing the extract through a 100-mesh filter cloth, and then the extract is filtered through a 200-mesh filter cloth. During the filtration process, the temperature of the extract is maintained at 40-50℃.

[0013] Preferably, in step S3, the Akkermansiamuciniphila powder added needs to be stored in a refrigerated environment at 2-8°C before use, and the feeding should be completed within 30 minutes after being taken out of the refrigerated environment. TRPV4 inhibitors need to be mixed evenly with an equal amount of excipients to form a premix before being added to the mixing system.

[0014] Preferably, in step S4, when preparing granules, a wet mixing granulator is used, purified water is used as a binder, the amount of binder added is 10-15% of the weight of the mixture, after granulation, the granules are dried in a hot air circulating drying oven at 45-50℃ until the moisture content of the granules is ≤6%, and then granulated through a 16-20 mesh sieve. After granulation, the fine powder that passes through an 80 mesh sieve is removed. When preparing tablets, a rotary tablet press is used, with a tableting pressure of 8-12 MPa, tablet hardness controlled at 3-5 kg, tablet disintegration time ≤30 minutes, and tablet weight variation limit ±3%.

[0015] This invention provides a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes and its preparation method. It possesses the following beneficial effects: 1. This invention employs a synergistic combination of traditional Chinese medicine compound, functional probiotics, and targeted inhibitors to achieve precise intervention across the entire chain of spermatogenesis disorders caused by high-altitude oxidative stress, providing comprehensive protection against oxidation, gut microbiota regulation, and immune damage. Compared to existing technologies using single antioxidants, hormone supplements, or traditional Chinese medicine compound formulas, this invention addresses the issues of limited efficacy, inability to cover multiple links of damage to the gut and testis axis, and high recurrence rates.

[0016] 2. This invention employs an optimized water extraction process combined with a precise temperature-controlled mixing preparation technique, which achieves the technical effect of fully preserving the active ingredients of traditional Chinese medicine and the activity of probiotics, while improving the stability and uniformity of the preparation. Compared with the existing extraction and preparation schemes that lack active ingredient protection design, this invention solves the problems of easy loss of effective ingredients, poor preparation stability, and difficulty in mass production.

[0017] 3. This invention adopts a multi-component synergistic and safe formulation design technology, which achieves both preventive and therapeutic effects and is suitable for various high-altitude exposure populations. Compared with existing technologies such as hormone-based Western medicines or single-target inhibitors, it solves the problems of obvious side effects, limited applicable scenarios, and inability to take both prevention and treatment into account. Attached Figure Description

[0018] Figure 1 This is a schematic flowchart illustrating the steps of preparing a composition for combating oxidative stress and spermatogenesis disorders caused by high altitude, according to the present invention. Detailed Implementation

[0019] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0020] This invention provides a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, comprising the following raw materials in parts by weight: 15-25 parts of wolfberry, 12-20 parts of polygonatum, 10-18 parts of dodder seed, 8-16 parts of angelica, 10-20 parts of astragalus, 10-18 parts of raspberry, 2-5 parts of Akkermansia muciniphila mycelium powder, and 0.01-0.05 parts of TRPV4 inhibitor.

[0021] It also includes excipients, which are selected from at least one of maltodextrin and magnesium stearate, and the weight of the excipients accounts for 10-20% of the total weight of the composition.

[0022] The TRPV4 inhibitor is HC-067047; Akkermansia muciniphila mycelium powder was obtained through aseptic preparation process; wolfberry, polygonatum, dodder seed, angelica, astragalus, and raspberry are all dried, mold-free, and impurity-free raw materials, among which wolfberry is dried fruit, polygonatum is dried product, dodder seed is dried seed, angelica is dried root, astragalus is dried root, and raspberry is dried fruit.

[0023] Specifically, the synergistic effects of the various raw materials construct a three-pronged intervention system that combats oxidative stress, regulates gut microbiota, and blocks testicular immune damage. The formulation of traditional Chinese medicine raw materials focuses on anti-oxidation and promoting spermatogenesis, while Akkermansiamuciniphila powder regulates gut microbiota balance and reduces the generation of abnormal metabolites. TRPV4 inhibitors precisely block the transduction of immune damage signals. The three work together to cover key links in the pathological chain of high spermatogenesis disorder.

[0024] Please see the appendix Figure 1 A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes includes the following steps: S1. Raw material pretreatment: The Chinese herbal raw materials are removed of impurities, washed, dried and then pulverized to obtain mixed Chinese herbal powder; In S1, the drying process uses hot air circulation drying at a temperature of 45-50℃ for 3-5 hours, and the moisture content of the dried Chinese herbal raw materials is ≤8%. The pulverizing process uses a universal pulverizer, and the pulverized Chinese medicine powder has a particle size of 80-100 mesh. After pulverizing, it is screened by a vibrating screen to remove coarse powder that does not meet the particle size requirements before mixing to obtain a uniformly mixed Chinese medicine powder. Specifically, the raw materials of Chinese medicine are manually cleaned to remove visible impurities and moldy parts. They are then rinsed three times with purified water and dried. After being crushed, they are sieved and mixed to ensure that the dissolution efficiency of the effective components of each raw material is consistent during the subsequent extraction process.

[0025] S2. Preparation of Chinese herbal extract: Add extraction solvent to the mixed Chinese herbal powder and soak it for multiple extractions. Combine the extracts and filter them. Concentrate the filtered extract under reduced pressure to obtain an extract paste. Then, vacuum dry the extract paste and pulverize it to obtain Chinese herbal extract powder. In S2, the extraction solvent is deionized water, and the amount of extraction solvent added is 10-15 times the volume of the mixed Chinese herbal powder. The soaking time is 2-3 hours, and the mixture is stirred once every 30 minutes for 1 minute each time. The extraction was performed twice using a gentle boiling process, with the first extraction lasting 2 hours and the second extraction lasting 1.5 hours. The vacuum concentration was achieved using a rotary evaporator under the following conditions: pressure -0.08 to -0.09 MPa and temperature 55-65°C. The resulting extract had a relative density of 1.15-1.20 at 60°C. Vacuum drying is performed using a vacuum drying oven. The drying conditions are a temperature of 50-55℃ and a pressure of -0.07 to -0.08 MPa. The moisture content of the dried extract is ≤5%. The dried extract is then pulverized using an ultra-micro pulverizer to obtain a traditional Chinese medicine extract powder with a particle size of 100-120 mesh. In S2, the extract is filtered in two stages: first, large particles of medicinal residue are removed by passing it through a 100-mesh filter cloth, and then it is filtered through a 200-mesh filter cloth. During the filtration process, the temperature of the extract is maintained at 40-50℃. Specifically, the water extraction process avoids organic solvent residue, two-stage filtration can fully remove impurities and reduce adsorption loss of active ingredients, and the parameter settings for vacuum concentration and vacuum drying can reduce energy consumption while maximizing the preservation of heat-sensitive active ingredients of traditional Chinese medicine.

[0026] S3. Mixing and blending: Under sterile conditions, first premix the Chinese herbal extract powder evenly, then add Akkermansiamuciniphila bacterial powder under preset temperature and humidity conditions, followed by TRPV4 inhibitor and excipients, and mix until the preset homogeneity standard is met. In S3, the sterile environment is constructed through ultraviolet disinfection for more than 30 minutes, with a cleanliness level of 100; the preset temperature and humidity conditions are temperature ≤25℃ and humidity ≤45%. The premixing process uses a three-dimensional motion mixer with a mixing speed of 200-300 r / min and a premixing time of 15-20 minutes; After adding Akkermansiamuciniphila powder, premix manually for 5 minutes, then add TRPV4 inhibitor and excipients, and transfer to a double helix cone mixer for mixing at a speed of 150-250 r / min for 25-35 minutes. The preset homogeneity standard is that the relative standard deviation (RSD) of the mixture is ≤3%; In S3, the Akkermansiamuciniphila powder added must be stored in a refrigerated environment at 2-8℃ before use, and the feeding must be completed within 30 minutes after being taken out of the refrigerated environment. TRPV4 inhibitors need to be mixed evenly with an equal amount of excipients to form a premix before being added to the mixing system; Specifically, staged mixing and temperature and humidity control can prevent the inactivation of Akkermansiamuciniphila powder, and the premixing treatment of TRPV4 inhibitor can solve the problem of uneven dispersion caused by its small dosage and easy agglomeration. The multi-stage mixing process ensures that each component is evenly distributed in the mixture.

[0027] S4. Formulation: The mixture is processed into granules, capsules or tablets using the corresponding process; In S4, the capsules are filled using a fully automatic capsule filling machine, with a specification of 0.3-0.5g / capsule. During the filling process, the fill weight difference is sampled and tested every 10 minutes, and the fill weight difference limit is ±5%. In S4, when preparing granules, a wet mixing granulator is used, with purified water as the binder. The amount of binder added is 10-15% of the weight of the mixture. After granulation, the granules are dried in a hot air circulating drying oven at 45-50℃ until the moisture content of the granules is ≤6%. Then, they are granulated through a 16-20 mesh sieve. After granulation, the fine powder that passes through an 80 mesh sieve is removed. When preparing tablets, a rotary tablet press is used, with a tableting pressure of 8-12 MPa, tablet hardness controlled at 3-5 kg, tablet disintegration time ≤30 minutes, and tablet weight variation limit ±3%.

[0028] Specifically, the process parameters for different dosage forms are optimized for the characteristics of the mixture. Granulation and fine powder removal steps can improve flowability, tablet hardness and disintegration time control can ensure dissolution efficiency after clinical administration, and random sampling inspection of capsule filler weight variation can ensure product quality uniformity.

[0029] The following is a description with reference to specific embodiments: Example 1: A composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, comprising the following raw materials in parts by weight: 15 parts of wolfberry, 12 parts of polygonatum, 10 parts of dodder seed, 8 parts of angelica, 10 parts of astragalus, 10 parts of raspberry, 2 parts of Akkermansia muciniphila powder, 0.01 parts of TRPV4 inhibitor HC-0670470, and maltodextrin as the excipient, with the excipient accounting for 10% of the total weight of the composition; Among them, Akkermansiamuciniphila mycelium powder is obtained through aseptic preparation process, wolfberry is dried fruit, polygonatum is dried product, dodder seed is dried seed, angelica is dried root, astragalus is dried root, and raspberry is dried fruit. All Chinese medicinal materials are dry, free from mold and impurities.

[0030] A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes includes the following steps: S1. Raw material pretreatment: The above-mentioned Chinese herbal raw materials are manually cleaned of impurities and then rinsed three times with purified water. They are then placed in a hot air circulating drying oven and dried at 45°C for 5 hours until the moisture content is ≤8%. After being pulverized by a universal pulverizer, they are screened through a 100-mesh vibrating screen to remove coarse powder. The powder is then mixed evenly to obtain mixed Chinese herbal powder.

[0031] S2. Preparation of Traditional Chinese Medicine Extract: Add 10 times the volume of deionized water to the mixed traditional Chinese medicine powder and soak for 2 hours. Stir for 1 minute every 30 minutes during soaking. Then extract twice by gentle boiling, the first time for 2 hours and the second time for 1.5 hours. Combine the extracts and filter through a 100-mesh filter cloth and a 200-mesh filter cloth. Keep the temperature of the extract at 40℃. Transfer the filtrate to a rotary evaporator and concentrate under reduced pressure at -0.09MPa and 55℃ to an extract with a relative density of 1.15 at 60℃. Then place the extract in a vacuum drying oven at 50℃ and -0.08MPa and dry until the moisture content is ≤5%. Use an ultra-micro pulverizer to pulverize to obtain a 100-mesh traditional Chinese medicine extract powder.

[0032] S3. Mixing and Blending: Transfer the Chinese herbal extract powder to a Class 100 cleanroom sterilized with ultraviolet light for at least 30 minutes. Premix it for 20 minutes at 200 rpm using a three-dimensional motion mixer. Control the temperature of the operating room to be ≤25℃ and the humidity to be ≤45%. Take out the Akkermansiamuciniphila bacterial powder stored at 2℃ and add it within 30 minutes. After manual premixing for 5 minutes, add the premix made of TRPV4 inhibitor and an equal amount of maltodextrin to the system. Transfer it to a double helix conical mixer and mix it at 150 rpm for 35 minutes. Detect the relative standard deviation (RSD) of the mixture to be ≤3%.

[0033] S4. Formulation: When preparing capsules, use a fully automatic capsule filling machine to fill to 0.3g / capsule, and sample and check the fill weight difference every 10 minutes, with a limit of ±5%; when preparing granules, add 10% purified water by weight to the mixture as a binder, wet mix and granulate, and dry with hot air circulation at 45℃ until the granule moisture content is ≤6%, granulate through a 16-mesh sieve and remove fine powder below 80 mesh; when preparing tablets, use a rotary tablet press to compress at a pressure of 8MPa, and control the tablet hardness to 3kg, disintegration time to ≤30 minutes, and tablet weight difference to ±3%.

[0034] Example 2: A composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, comprising the following raw materials in parts by weight: 20 parts of wolfberry, 16 parts of polygonatum, 14 parts of dodder seed, 12 parts of angelica, 15 parts of astragalus, 14 parts of raspberry, 3.5 parts of Akkermansia muciniphila powder, and 0.03 parts of TRPV4 inhibitor HC-067047. The excipient is a mixture of maltodextrin and magnesium stearate in a weight ratio of 1:1, and the excipient accounts for 15% of the total weight of the composition. Among them, Akkermansiamuciniphila mycelium powder is obtained through aseptic preparation process, wolfberry is dried fruit, polygonatum is dried product, dodder seed is dried seed, angelica is dried root, astragalus is dried root, and raspberry is dried fruit. All Chinese medicinal materials are dry, free from mold and impurities.

[0035] A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes includes the following steps: S1. Raw material pretreatment: The above-mentioned Chinese herbal raw materials are manually cleaned of impurities and then rinsed three times with purified water. They are then placed in a hot air circulating drying oven and dried at 48°C for 4 hours until the moisture content is ≤8%. After being pulverized by a universal pulverizer, they are screened through a 100-mesh vibrating screen to remove coarse powder. The powder is then mixed evenly to obtain mixed Chinese herbal powder.

[0036] S2. Preparation of Traditional Chinese Medicine Extract: Add 12 times the volume of deionized water to the mixed traditional Chinese medicine powder and soak for 2.5 hours. Stir for 1 minute every 30 minutes during soaking. Then extract twice by gentle boiling, the first time for 2 hours and the second time for 1.5 hours. Combine the extracts and filter through a 100-mesh filter cloth and a 200-mesh filter cloth. Keep the temperature of the extract at 45℃. Transfer the filtrate to a rotary evaporator and concentrate under reduced pressure at -0.085MPa and 60℃ to an extract with a relative density of 1.18 at 60℃. Then place the extract in a vacuum drying oven at 52℃ and -0.075MPa and dry until the moisture content is ≤5%. Use an ultra-micro pulverizer to obtain a 110-mesh traditional Chinese medicine extract powder.

[0037] S3. Mixing and Blending: Transfer the Chinese herbal extract powder to a Class 100 cleanroom sterilized with ultraviolet light for at least 30 minutes. Premix the powder for 18 minutes at 250 rpm using a three-dimensional motion mixer. Control the temperature of the operating room to be ≤25℃ and the humidity to be ≤45%. Take out the Akkermansiamuciniphila bacterial powder stored at 5℃ and add it within 30 minutes. After manual premixing for 5 minutes, add the premix made of TRPV4 inhibitor and an equal amount of excipients to the system. Transfer the mixture to a double helix conical mixer and mix at 200 rpm for 30 minutes. Detect the relative standard deviation (RSD) of the mixture to be ≤3%.

[0038] S4. Formulation: When preparing capsules, use a fully automatic capsule filling machine to fill to 0.4g / capsule, and sample and check the fill weight difference every 10 minutes, with a limit of ±5%; when preparing granules, add 12% purified water by weight to the mixture as a binder, wet mix and granulate, and dry with hot air circulation at 48℃ until the granule moisture content is ≤6%, granulate through an 18-mesh sieve and remove fine powder below 80 mesh; when preparing tablets, use a rotary tablet press to compress at a pressure of 10MPa, and control the tablet hardness to 4kg, disintegration time to ≤30 minutes, and tablet weight difference to ±3%.

[0039] Example 3: A composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, comprising the following raw materials in parts by weight: 25 parts of wolfberry, 20 parts of polygonatum, 18 parts of dodder seed, 16 parts of angelica, 20 parts of astragalus, 18 parts of raspberry, 5 parts of Akkermansia muciniphila mycelium powder, and 0.05 parts of TRPV4 inhibitor HC-0670470. The excipient is magnesium stearate, and the excipient accounts for 20% of the total weight of the composition. Among them, Akkermansiamuciniphila mycelium powder is obtained through aseptic preparation process, wolfberry is dried fruit, polygonatum is dried product, dodder seed is dried seed, angelica is dried root, astragalus is dried root, and raspberry is dried fruit. All Chinese medicinal materials are dry, free from mold and impurities.

[0040] A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes includes the following steps: S1. Raw material pretreatment: The above-mentioned Chinese herbal raw materials are manually cleaned of impurities and then rinsed three times with purified water. They are then placed in a hot air circulating drying oven and dried at 50°C for 3 hours until the moisture content is ≤8%. After being pulverized by a universal pulverizer, they are screened through a 100-mesh vibrating screen to remove coarse powder. The powder is then mixed evenly to obtain mixed Chinese herbal powder.

[0041] S2. Preparation of Traditional Chinese Medicine Extract: Add 15 times the volume of deionized water to the mixed traditional Chinese medicine powder and soak for 3 hours. Stir for 1 minute every 30 minutes during soaking. Then extract twice by gentle boiling, the first time for 2 hours and the second time for 1.5 hours. Combine the extracts and filter through a 100-mesh filter cloth and a 200-mesh filter cloth. Keep the temperature of the extract at 50℃. Transfer the filtrate to a rotary evaporator and concentrate under reduced pressure at -0.08MPa and 65℃ to an extract with a relative density of 1.20 at 60℃. Then place the extract in a vacuum drying oven at 55℃ and -0.07MPa and dry until the moisture content is ≤5%. Use an ultra-fine pulverizer to obtain a 120-mesh traditional Chinese medicine extract powder.

[0042] S3. Mixing and Blending: Transfer the Chinese herbal extract powder to a Class 100 cleanroom sterilized with ultraviolet light for at least 30 minutes. Premix it for 15 minutes at 300 rpm using a three-dimensional motion mixer. Control the temperature of the operating room to be ≤25℃ and the humidity to be ≤45%. Take out the Akkermansiamuciniphila bacterial powder stored at 8℃ and add it within 30 minutes. After manual premixing for 5 minutes, add the premixing agent made of TRPV4 inhibitor and an equal amount of magnesium stearate to the system. Transfer it to a double helix conical mixer and mix it at 250 rpm for 25 minutes. Detect the relative standard deviation (RSD) of the mixture to be ≤3%.

[0043] S4. Formulation: When preparing capsules, use a fully automatic capsule filling machine to fill to 0.5g / capsule, and sample and check the fill weight difference every 10 minutes, with a limit of ±5%; when preparing granules, add 15% purified water by weight to the mixture as a binder, wet mix and granulate, and dry with hot air circulation at 50℃ until the granule moisture content is ≤6%, sieve through a 20-mesh sieve to remove fine powder below 80 mesh; when preparing tablets, use a rotary tablet press to compress at a pressure of 12MPa, controlling the tablet hardness to 5kg, disintegration time to ≤30 minutes, and tablet weight difference to ±3%.

[0044] Comparative Example 1: Unlike Example 2, the preparation process of the Chinese herbal extract was changed to ethanol reflux extraction. Specifically, 12 times the volume of 70% ethanol was added to the mixed Chinese herbal powder, and after soaking for 2.5 hours, the mixture was refluxed twice, for 2 hours the first time and 1.5 hours the second time. The combined extracts were filtered through a 100-mesh filter cloth and a 200-mesh filter cloth, maintaining the temperature of the extract at 45°C. The filtrate was then transferred to a rotary evaporator and concentrated under reduced pressure at -0.085 MPa and 60°C to a paste with a relative density of 1.18 at 60°C. The paste was then placed in a vacuum drying oven and dried at 52°C and -0.075 MPa until the moisture content was ≤5%. The paste was then pulverized using an ultrafine pulverizer to obtain a 110-mesh Chinese herbal extract powder. All other process parameters were the same as in Example 2.

[0045] Comparative Example 2: Unlike Example 2, the premixing step of the Chinese herbal extract powder was omitted in the mixing and preparation process. The Chinese herbal extract powder was directly transferred to a Class 100 cleanroom sterilized with ultraviolet light for more than 30 minutes. The temperature of the room was controlled at ≤25℃ and the humidity at ≤45%. Akkermansiamuciniphila bacterial powder stored at 5℃ was taken out and added within 30 minutes. After manual premixing for 5 minutes, a premix made of TRPV4 inhibitor and an equal amount of excipients was added. The mixture was then transferred to a double helix conical mixer and mixed at 200 r / min for 30 minutes. The relative standard deviation (RSD) of the mixture was ≤3%. All other process parameters were the same as in Example 2.

[0046] Comparative Example 3: Unlike Example 2, after the preparation of the Chinese herbal extract was completed, the Akkermansiamuciniphila bacterial powder was mixed with the Chinese herbal extract and dried together in a vacuum drying oven at 52°C and -0.075MPa. After drying until the moisture content was ≤5%, the mixture was pulverized to obtain a mixed powder. In subsequent mixing and formulation, TRPV4 inhibitors and excipients were directly added, and the mixture was transferred to a double helical conical mixer and mixed at 200r / min for 30 minutes. The relative standard deviation (RSD) of the mixture was ≤3%, and the remaining process parameters were the same as in Example 2.

[0047] Comparative Example 4: Unlike Example 2, the granulation step was omitted during formulation. For capsule preparation, the mixture was directly filled to 0.4g / capsule using a fully automatic capsule filling machine. For tablet preparation, the mixture was directly added to a rotary tablet press and compressed at 10MPa pressure. The tablet hardness was controlled at 4kg, the disintegration time was ≤30 minutes, and the tablet weight difference was ±3%. All other process parameters were the same as in Example 2.

[0048] Comparative Example 5: Unlike Example 2, the temperature and humidity of the aseptic operating room were not controlled during the mixing and preparation process. Akkermansiamuciniphila powder was added directly under normal temperature and humidity conditions. The subsequent manual premixing and double helix cone mixer mixing steps were all carried out under normal temperature and humidity conditions. All other process parameters were the same as in Example 2.

[0049] Table 1,

[0050] Based on the differences between Examples 1-3 and Comparative Examples 1-5 and the performance test data table, it can be seen that the key factors in this invention, such as the synergistic compatibility of the ternary components, the optimization of the water extraction process of traditional Chinese medicine, the temperature and humidity control of probiotics, the homogenization treatment, and the adaptation of the formulation molding process, have a significant impact on the retention rate of active ingredients, the survival rate of probiotics, the stability and homogeneity of the formulation, and the comprehensive efficacy against high-altitude oxidative stress and spermatogenesis disorders. Moreover, there are good effects of synergistic enhancement of components, complementary enhancement of processes, and stable adaptation of performance among the various factors.

[0051] Comparative Example 1, by changing the water extraction process of traditional Chinese medicine to ethanol reflux extraction, resulted in a significant decrease in the retention rates of the core active ingredients, Lycium barbarum polysaccharide and Polygonatum sibiricum polysaccharide, which dropped to 76.2% and 74.5% respectively, a decrease of 19.1% and 21.3% compared to 95.3% and 94.6% in Example 2. After 6 months of storage at room temperature, the total retention rate of active ingredients was only 68.4%, a decrease of 24.3% compared to Example 2, making it the group with the worst retention of active ingredients. Comparative Example 2, by omitting the premixing step of the traditional Chinese medicine extract, caused the RSD of the mixture homogeneity to rise to 4.2%, an increase of 180% compared to 1.5% in Example 2. The difference in capsule weight also widened to ±3.8%, an increase of 81% compared to Example 2. Uneven component dispersion affected the overall efficacy. Comparative Example 3, by simultaneously drying probiotics and traditional Chinese medicine extract at high temperature, resulted in Akkermansiamucin... The survival rate of iphila live bacteria was only 38.6%, a decrease of 58.4% compared to 92.7% in Example 2, indicating that its intestinal microecological regulation function was basically ineffective. After 6 months of storage, the total retention rate of active ingredients was only 51.3%, and the overall performance declined across the board. In Comparative Example 4, the granulation step was omitted during formulation, and capsules or tablets were directly filled or compressed, resulting in a decrease in tablet hardness to 3.2 kg, a decrease of 20% compared to Example 2. The disintegration time was extended to 28.7 min, an increase of 54.3% compared to Example 2. The capsule dosage varied by ±4.5%, indicating a significant lack of formulation process adaptability. In Comparative Example 5, temperature and humidity were not controlled during mixing and preparation, resulting in a decrease in the survival rate of probiotic live bacteria to 65.8%, a decrease of 29% compared to Example 2. After 6 months of storage, the total retention rate of active ingredients was only 78.2%, a decrease of 13.4% compared to Example 2, which also affected the long-term stability of the formulation.

[0052] In the examples, the composition for combating high-altitude oxidative stress and spermatogenesis disorders exhibited excellent core performance indicators: Lycium barbarum polysaccharide retention rate 87.3%-95.3%, Polygonatum sibiricum polysaccharide retention rate 87.3%-94.6%, Akkermansia muciniphila live bacteria survival rate 85.2%-92.7%, mixture homogeneity RSD 1.5%-2.8%, total active ingredient retention rate after 6 months of room temperature storage 82.5%-90.3%, capsule weight variation ±2.1%-±4.2%, tablet hardness 3.0-5.0 kg, disintegration time 18.6-25.3 min, and in vitro DPPH free radical scavenging rate 76.8%-89.5%. Among them, Examples 1 and 3 are slightly inferior to Example 2 in performance due to their formulation and process parameters being at the boundary of the set range, but they still maintain good overall advantages. Example 1 is suitable for low-cost large-scale production scenarios. By optimizing the amount of raw materials and process parameters, the cost is reduced by 8% compared with Example 2, while maintaining the retention rate of core components and the survival rate of probiotics, meeting the basic prevention and treatment needs. Example 3 is designed for high-dose enhancement scenarios. By increasing the amount of Chinese medicine raw materials and probiotics, it enhances the antioxidant and intestinal regulation effects, and the in vitro DPPH free radical scavenging rate reaches 88.9%, which is suitable for the deep protection needs of people exposed to high altitudes for a long time.

[0053] Example 2 achieves an optimal balance between performance and cost, with a 95.3% retention rate of Lycium barbarum polysaccharides, a 94.6% retention rate of Polygonatum sibiricum polysaccharides, a 92.7% survival rate of live probiotics, a 1.5% RSD for the homogeneity of the mixture, a 90.3% total retention rate of active ingredients after 6 months of storage, a tablet hardness of 4.0 kg, a disintegration time of 18.6 min, and an in vitro DPPH free radical scavenging rate of 89.5%. Furthermore, the preparation process exhibits strong compatibility among various process parameters, sufficient synergistic effects of the components, high formulation stability, and a short production cycle, facilitating large-scale production. Its cost-effectiveness is significant, reducing costs by 10% compared to Example 3 and improving overall efficacy and stability by more than 40% compared to traditional single-component formulations.

[0054] In Examples 1-3, the core processes and formulations form a deep synergistic effect: the proportion of Chinese herbal raw materials and the water extraction parameters such as soaking time, number of extractions, and concentration and drying conditions are precisely matched to ensure the efficient retention of active ingredients; the storage conditions of probiotics, the timing of feeding, and the temperature and humidity of the mixing environment are coordinated to ensure the survival rate of live bacteria; the premixing parameters of Chinese herbal extracts, manual premixing of probiotics, and final mixing parameters of the double helix conical mixer are matched to achieve the uniformity of the mixture; the granulation parameters, filling / tableting parameters, and material characteristics of the formulation forming process are matched to ensure the stability of the formulation morphology and performance. Example 2 optimizes the formula ratio of 20 parts wolfberry, 16 parts polygonatum, 14 parts dodder seed, 12 parts angelica, 15 parts astragalus, 14 parts raspberry, 3.5 parts Akkermansia muciniphila powder, and 0.03 parts TRPV4 inhibitor. The extraction parameters include soaking in 12 times the volume of deionized water for 2.5 hours, extraction twice by gentle boiling, concentration under reduced pressure to a relative density of 1.18, vacuum drying at 52℃, premixing at 250 rpm for 18 minutes using a three-dimensional motion mixer, final mixing at 200 rpm for 30 minutes using a double-helix conical mixer, maintaining a temperature and humidity of ≤25℃ / 45%, and adding 12% binder, drying at 48℃, and granulation at 18 mesh. This optimizes the formula ratio, maximizing synergistic advantages and demonstrating the scientific validity and rationality of the formula range and process parameters of this invention.

[0055] Comparative Examples 1-5 suffered from the breakdown of the overall synergistic chain due to the absence of a single process step or changes in the formulation ratio: Comparative Example 1 replaced the extraction solvent and process, directly causing the loss of active ingredients; Comparative Example 2 eliminated the premixing step, destroying the basis of component homogeneity; Comparative Example 3 changed the timing of probiotic addition and drying conditions, resulting in the inactivation of live bacteria; Comparative Example 4 simplified the formulation molding process, affecting the mechanical properties and stability of the formulation; Comparative Example 5 neglected the control of temperature and humidity in the mixing environment, damaging the activity of probiotics and long-term storage stability. All of these examples demonstrate the irreplaceable nature of the key formulation ratios and process steps of this invention.

[0056] In summary, this invention, through the synergistic design of a three-component formula combining traditional Chinese medicine compound, functional probiotics, and targeted inhibitors, and the synergistic innovation of processes such as optimized water extraction of traditional Chinese medicine, precise protection of probiotics, homogenization of mixing, and formulation adaptation, solves the technical problems of existing technologies, such as limited efficacy of single-target intervention, easy loss of active ingredients, low survival rate of probiotics, and poor formulation stability. It significantly improves the comprehensive efficacy and clinical applicability of anti-high-altitude oxidative stress spermatogenesis disorders. Example 2 achieves an optimal balance between performance, cost, and production efficiency, making it suitable for large-scale promotion. Examples 1-3 cover multiple application scenarios, from low-cost general-purpose to high-dose intensive formulations, fully adapting to the prevention and treatment needs of different high-altitude exposed populations, providing a replicable and scalable technical solution for the industrial production of anti-high-altitude spermatogenesis disorder preparations.

[0057] Retention rate of Lycium barbarum polysaccharides: Referencing GB / T22244-2018 "Determination of Lycium barbarum Polysaccharides in Health Foods". High performance liquid chromatography (HPLC) was used, with glucose as the reference standard. An amino column was selected, the mobile phase was acetonitrile-water (70:30), and the detection wavelength was set to 360 nm. The retention rate was calculated by determining the content of Lycium barbarum polysaccharides in the sample.

[0058] Retention rate of Polygonatum polysaccharides: Refer to GB / T39564-2020 "Determination of Polygonatum polysaccharides". The phenol-sulfuric acid colorimetric method was used, with glucose as a reference standard. The absorbance was measured at a wavelength of 490 nm. After plotting a standard curve, the content of Polygonatum polysaccharides in the sample was calculated and the retention rate was converted.

[0059] Viable bacterial survival rate of Akkermansia muciniphila: Refer to GB4789.35-2016 "National Food Safety Standard - Microbiological Examination of Food - Examination of Lactic Acid Bacteria". The plate count method was used. Samples were serially diluted and inoculated onto modified MRS medium, incubated anaerobically at 37°C for 48 hours, and the number of viable colonies was counted to calculate the survival rate.

[0060] Mixture homogeneity: Refer to GB / T19077-2008 "Particle size analysis by laser diffraction". Three batches of samples were selected, and the content of key components (goji berry polysaccharide and polygonatum polysaccharide) was detected, and the relative standard deviation (RSD) was calculated. At the same time, the uniformity of particle size distribution was detected by laser diffraction to comprehensively characterize the homogeneity of the mixture.

[0061] Total retention rate of active ingredients after 6 months of storage at room temperature: Refer to GB / T23788-2009 "Technical Specifications for Inspection and Evaluation of Health Foods". After sealing the samples, store them at room temperature (25℃±2℃, humidity 60%±5%) for 6 months. Determine the content according to the corresponding detection methods for Lycium barbarum polysaccharides and Polygonatum sibiricum polysaccharides, and calculate the total retention rate of active ingredients.

[0062] Capsule fill weight variation: Refer to GB13052-2008 "General Rules for Capsules". Randomly select 20 capsules, accurately weigh the contents of each capsule, calculate the average fill weight and the difference in fill weight of each capsule, and determine the limit of fill weight variation.

[0063] Angle of repose for granules: Refer to GB / T11986-2008 "Determination of Angle of Repose of Surfactant Powders and Particles". Using the fixed funnel method, the granules are allowed to fall freely through the funnel onto a horizontal table, forming a cone. The angle between the generatrix of the cone and the horizontal plane is measured; this is the angle of repose.

[0064] Tablet hardness: Refer to GB / T28728-2012 "Methods for Determination of Tablet Hardness and Friability". A tablet hardness tester was used to randomly select 10 samples, and their hardness was measured. The average value was taken as the tablet hardness test result.

[0065] Tablet disintegration time: Refer to Appendix 6 of GB15980-1995 "Regulations on the Management of Drug Packaging, Labeling and Instructions". Use a disintegration time tester to place the tablets in purified water at 37℃±1℃ and record the time it takes for the tablets to completely disintegrate and pass through a sieve.

[0066] In vitro DPPH free radical scavenging rate: Refer to GB / T31740-2015 "Determination of Antioxidant Capacity of Tea Products - DPPH Method". The DPPH colorimetric method was used. The sample extract was mixed with the DPPH free radical solution, and the mixture was reacted at room temperature in the dark for 30 minutes. The absorbance was measured at a wavelength of 517 nm, and the free radical scavenging rate was calculated.

[0067] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, characterized in that, The ingredients include the following parts by weight: 15-25 parts wolfberry, 12-20 parts polygonatum, 10-18 parts dodder seed, 8-16 parts angelica, 10-20 parts astragalus, 10-18 parts raspberry, 2-5 parts Akkermansiamuciniphila mycelium powder, and 0.01-0.05 parts TRPV4 inhibitor.

2. The composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes according to claim 1, characterized in that: It also includes excipients selected from at least one of maltodextrin and magnesium stearate, wherein the weight of the excipients accounts for 10-20% of the total weight of the composition.

3. The composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes according to claim 1, characterized in that: The TRPV4 inhibitor is HC-067047; the Akkermansiamuciniphila mycelium powder is obtained through an aseptic preparation process; the wolfberry, polygonatum, dodder seed, angelica, astragalus, and raspberry are all dried, mold-free, and impurity-free raw materials, wherein wolfberry is dried fruit, polygonatum is dried product, dodder seed is dried seed, angelica is dried root, astragalus is dried root, and raspberry is dried fruit.

4. A method for preparing a composition for combating spermatogenesis disorders caused by oxidative stress at high altitudes, characterized in that: Includes the following steps: S1. Raw material pretreatment: The Chinese herbal raw materials are removed of impurities, washed, dried and then pulverized to obtain mixed Chinese herbal powder; S2. Preparation of Chinese herbal extract: Add extraction solvent to the mixed Chinese herbal powder and soak it for multiple extractions. Combine the extracts and filter them. Concentrate the filtered extract under reduced pressure to obtain an extract paste. Then, vacuum dry the extract paste and pulverize it to obtain Chinese herbal extract powder. S3. Mixing and blending: Under sterile conditions, first premix the Chinese herbal extract powder evenly, then add Akkermansiamuciniphila bacterial powder under preset temperature and humidity conditions, followed by TRPV4 inhibitor and excipients, and mix until the preset homogeneity standard is met. S4. Formulation: The mixture is processed into granules, capsules or tablets using the corresponding process.

5. A method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In step S1, the drying process employs a hot air circulation drying method, with a drying temperature of 45-50℃ and a drying time of 3-5 hours. The moisture content of the dried Chinese herbal raw materials is ≤8%. The powder is pulverized using a universal pulverizer. The resulting powder has a particle size of 80-100 mesh. After pulverization, the powder is screened through a vibrating screen to remove coarse powder that does not meet the particle size requirements before mixing to obtain a uniformly mixed powder.

6. The method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In step S2, the extraction solvent is deionized water, and the amount of extraction solvent added is 10-15 times the volume of the mixed Chinese herbal powder. The soaking time is 2-3 hours, and the mixture is stirred once every 30 minutes for 1 minute each time. The extraction was performed twice using a gentle boiling process, with the first extraction lasting 2 hours and the second extraction lasting 1.5 hours. The vacuum concentration was achieved using a rotary evaporator under the following conditions: pressure -0.08 to -0.09 MPa and temperature 55-65°C. The resulting extract had a relative density of 1.15-1.20 at 60°C. Vacuum drying is performed using a vacuum drying oven under the following conditions: temperature 50-55℃, pressure -0.07~-0.08MPa. The moisture content of the dried extract is ≤5%. The dried extract is then pulverized using an ultra-micro pulverizer to obtain a traditional Chinese medicine extract powder with a particle size of 100-120 mesh.

7. A method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In S3, a sterile environment is created by ultraviolet disinfection for more than 30 minutes, with a cleanliness level of Class 100; the preset temperature and humidity conditions are temperature ≤25℃ and humidity ≤45%. The premixing process uses a three-dimensional motion mixer with a mixing speed of 200-300 r / min and a premixing time of 15-20 minutes; After adding Akkermansiamuciniphila powder, premix manually for 5 minutes, then add TRPV4 inhibitor and excipients, and transfer to a double helix cone mixer for mixing at a speed of 150-250 r / min for 25-35 minutes. The preset homogeneity standard is that the relative standard deviation (RSD) of the mixture is ≤3%; In step S4, the capsules are filled using a fully automatic capsule filling machine with a specification of 0.3-0.5g / capsule. During the filling process, the filling volume difference is sampled and tested every 10 minutes, and the filling volume difference limit is ±5%.

8. A method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In S2, the extract is filtered in two stages: first, large particles of medicinal residue are removed by passing the extract through a 100-mesh filter cloth, and then it is filtered through a 200-mesh filter cloth. During the filtration process, the temperature of the extract is maintained at 40-50℃.

9. A method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In S3, the Akkermansiamuciniphila powder added must be stored in a refrigerated environment at 2-8℃ before use, and the feeding must be completed within 30 minutes after being taken out of the refrigerated environment. TRPV4 inhibitors need to be mixed evenly with an equal amount of excipients to form a premix before being added to the mixing system.

10. A method for preparing a composition for combating high-altitude oxidative stress and spermatogenesis disorders according to claim 4, characterized in that: In S4, when preparing granules, a wet mixing granulator is used, purified water is used as a binder, and the amount of binder added is 10-15% of the weight of the mixture. After granulation, the granules are dried in a hot air circulating drying oven at 45-50℃ until the moisture content of the granules is ≤6%, and then granulated through a 16-20 mesh sieve. After granulation, the fine powder that passes through an 80 mesh sieve is removed. When preparing tablets, a rotary tablet press is used, with a tableting pressure of 8-12 MPa, tablet hardness controlled at 3-5 kg, tablet disintegration time ≤30 minutes, and tablet weight variation limit ±3%.

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