Formula for tonifying kidney, strengthening tendons and supporting yang and preparation method thereof

Abstract

The invention relates to the technical field of traditional Chinese medicine preparations, in particular to a kidney-tonifying, tendon-strengthening and yang-supporting formula and a preparation method thereof.The kidney-tonifying, tendon-strengthening and yang-supporting formula is prepared from, by weight, 50-70 g of obscured homalomena rhizome, 50-70 g of salted morinda officinalis, 40-60 g of fructus alpiniae oxyphyllae, 40-60 g of beautiful millettia roots, 30-50 g of gordon euryale seeds, 30-50 g of Chinese yam, 20-40 g of medicinal cyathula roots, 10-20 g of cinnamon, 10-20 g of citrus chachiensis hortorum and 400-500 g of honey. The compatibility of the whole formula embodies the compatibility characteristics of warming yang without dryness, tonifying without stagnation and nourishing without greasiness, and is especially suitable for conditioning and treating male climacteric syndrome and delayed hypogonadism in humid and hot climatic environments in Lingnan regions.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine preparation technology, specifically to a kidney-tonifying, muscle-strengthening, and yang-boosting formula and its preparation method. This invention is mainly applied to the prevention and treatment of male menopausal syndrome and late-onset hypogonadism, and is particularly suitable for the health conditioning needs of men in the humid and hot climate of the Lingnan region. Background Technology

[0002] Male menopause syndrome refers to a series of physical and psychological symptoms experienced by men transitioning from middle age to old age due to a gradual decline in androgen levels. These symptoms primarily manifest as decreased sexual function, reduced physical performance, and mood changes. Late-onset hypogonadism (LOH) refers to a clinical syndrome in adult men caused by decreased testicular function, resulting in serum testosterone levels below the normal range. Clinical manifestations include decreased libido, erectile dysfunction, lack of energy, decreased muscle strength, and decreased bone density. With the accelerating aging of the population, the incidence of male menopause syndrome and LOH has been rising annually, becoming a significant health issue affecting the quality of life for middle-aged and elderly men.

[0003] Currently, the main clinical treatment for this type of disease is androgen replacement therapy. While it can quickly relieve symptoms, long-term use carries adverse effects such as increased risk of prostate cancer, inducing cardiovascular events, and affecting lipid metabolism. Traditional Chinese medicine (TCM) has unique advantages in treating male menopausal syndrome and late-onset hypogonadism. It addresses both the symptoms and the root cause through holistic regulation, with fewer adverse reactions. TCM believes that the pathogenesis of male menopausal syndrome and late-onset hypogonadism is mainly due to kidney yang deficiency, insufficient essence and blood, and malnourishment of the tendons and bones. Treatment should focus on warming and tonifying kidney yang, replenishing essence and marrow, and strengthening tendons and bones.

[0004] Chinese patent application CN115006503A discloses a "Golden Bull Kidney Tonifying Paste" and its preparation method. The patented formula includes eight traditional Chinese medicinal herbs: Rosa laevigata, Achyranthes bidentata, red ginseng rootlets, Schisandra chinensis, Euryale ferox, Alpinia oxyphylla, Rubus idaeus, and Acorus tatarinowii. It is prepared as a paste using traditional methods of decoction, alcohol precipitation, and honey refining. It is primarily used to treat symptoms such as frequent urination, lower back pain, leg pain, weakness in the limbs, chronic cough, and insomnia caused by lung and kidney qi deficiency. While this patented formula focuses on nourishing yin and astringing, and has some effect on improving symptoms like frequent urination, its composition emphasizes astringency and lacks warming and tonifying yang herbs. Therefore, it is not highly targeted at the core symptoms of male menopause syndrome and late-onset hypogonadism, such as impotence, premature ejaculation, and low libido. Furthermore, the traditional decoction method used in this patented preparation process involves high extraction temperatures and long extraction times, which can easily lead to the loss of heat-sensitive components such as volatile oils, affecting product quality and efficacy.

[0005] The Lingnan region has a hot and humid climate, and men with male menopausal syndrome and late-onset hypogonadism often exhibit a constitution characterized by internal damp-heat. Therefore, while tonifying kidney yang, it is important to regulate qi and resolve dampness, avoiding greasy and cloying foods that may hinder digestion. The Lingnan region is rich in various authentic medicinal herbs, such as salt-processed Morinda officinalis, dried tangerine peel, Achyranthes bidentata, and cinnamon. These herbs not only suit the constitution of the Lingnan population but also possess excellent kidney-tonifying, yang-boosting, and bone-strengthening effects. Developing traditional Chinese medicine pastes suitable for treating male menopausal syndrome and late-onset hypogonadism in the Lingnan region by fully utilizing these authentic medicinal resources has significant clinical and social value.

[0006] Therefore, in view of the shortcomings of existing technologies, there is an urgent need to develop a traditional Chinese medicine ointment with a reasonable formula, scientific process, controllable quality, and definite curative effect for the prevention and treatment of male menopausal syndrome and late-onset hypogonadism. Summary of the Invention

[0007] The purpose of this invention is to provide a kidney-tonifying, muscle-strengthening, and yang-boosting formula and its preparation method, to overcome the shortcomings of existing technologies. The ointment described in this invention has the effects of tonifying the kidneys and boosting yang, strengthening muscles and bones, and is mainly used to treat symptoms such as lower back and knee pain, impotence, premature ejaculation, low libido, fatigue, insomnia, forgetfulness, and osteoporosis caused by male menopausal syndrome and late-onset hypogonadism.

[0008] To achieve the above objectives, the technical solution adopted by the present invention is to provide a kidney-tonifying, muscle-strengthening, and yang-boosting formula, which is prepared from the following raw materials in parts by weight: 50g-70g of *Homalomena occulta*, 50g-70g of *Morinda officinalis* (salted), 40g-60g of *Alpinia oxyphylla*, 40g-60g of *Achyranthes bidentata*, 30g-50g of *Euryale ferox*, 30g-50g of *Dioscorea opposita*, 20g-40g of *Achyranthes bidentata*, 10g-20g of cinnamon, 10g-20g of dried tangerine peel, and 400g-500g of honey, which are boiled into a paste of 1000g.

[0009] In the preferred technical solution, the raw materials of the above-mentioned kidney-tonifying, muscle-strengthening, and yang-boosting formula are composed of the following ingredients by weight: 60g of *Homalomena occulta*, 60g of *Morinda officinalis* (salted), 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, 30g of *Achyranthes bidentata*, 15g of cinnamon, 15g of dried tangerine peel, and 450g of honey, which are boiled into a paste of 1000g.

[0010] This invention also provides a method for preparing the above-mentioned kidney-tonifying, muscle-strengthening, and yang-boosting formula, which includes the following steps:

[0011] First, the raw materials are pre-treated. The nine medicinal materials, namely, *Homalomena occulta*, *Morinda officinalis*, *Alpinia oxyphylla*, *Achyranthes bidentata*, *Euryale ferox*, *Dioscorea opposita*, *Achyranthes bidentata*, cinnamon, and dried tangerine peel, are weighed according to their weight proportions and then pulverized through an 80-mesh sieve. Among them, *Homalomena occulta*, cinnamon, and dried tangerine peel are pulverized at a low temperature of 5℃-10℃. The pulverized medicinal materials are then mixed evenly to obtain a mixed medicinal powder.

[0012] Enzymatic hydrolysis-assisted extraction was then performed. 360g of mixed medicinal powder was taken, 2500mL of distilled water was added, and cellulase with an enzyme activity of over 10000U / g was added at a dosage of 0.3% of the weight of the medicinal material. The pH was adjusted to 4.5-5.0, and the mixture was enzymatically hydrolyzed in a constant temperature water bath at 45℃ for 1.5h, with stirring every 30min during the hydrolysis process.

[0013] Next, ultrasonic extraction was performed. The enzymatic hydrolysate was transferred to an ultrasonic extraction device and ultrasonically extracted for 40 minutes at an ultrasonic power of 400W and a frequency of 40kHz.

[0014] Then, decoction extraction is carried out. The extract is heated to boiling and then simmered at 95℃-100℃ for 1.5 hours. The evaporated water is replenished in time to keep the liquid level stable. After decoction, the liquid is filtered through four layers of gauze and collected. The residue is added to 1800mL of distilled water and decocted for a second time for 1.5 hours. The residue is then filtered and collected. The two filtrates are combined to obtain 3500mL-4000mL of medicinal liquid.

[0015] The solution was then clarified by double filtration. After cooling the solution to room temperature, it was refrigerated at 4℃-8℃ for 12 hours. After being taken out, it was first filtered through eight layers of gauze, and then finely filtered through a microporous membrane with a pore size of 0.45μm to obtain a clarified solution.

[0016] Then, alcohol precipitation purification was carried out. 95% ethanol was slowly added to the clear drug solution while stirring. The ethanol volume content was adjusted to 60%. After standing at 4℃-8℃ for 24 hours, the solution was filtered through eight layers of gauze, the precipitate was discarded, and the filtrate was collected.

[0017] The filtrate was then concentrated under vacuum conditions of -0.08 MPa to -0.09 MPa and 55℃ to 65℃. Ethanol was first recovered until no alcohol odor was detected, and then the filtrate was further concentrated to a thick paste with a relative density of 1.30 to 1.35. The relative density was measured at 60℃.

[0018] Next, refine the honey to form a paste. Take 450g of honey and heat it to 110℃-115℃ for 5-8 minutes to refine the honey. After cooling to 80℃, add the concentrated paste and mix thoroughly. Continue heating and refining the paste at 80℃-85℃ while stirring constantly. Use the droplet bead test to check the consistency. Refine the paste to a total weight of 1000g.

[0019] Then, cooling and filling are carried out. The paste is quickly filled into sterilized containers while it is still hot at 60℃-70℃, with each bottle containing 250g or 500g. After filling, the containers are immediately sealed and inverted to cool to room temperature.

[0020] Finally, quality inspection is carried out, including checking the appearance, moisture content, microbial limits, heavy metals and harmful elements, and content of indicator components. After passing the inspection, the finished product is packaged.

[0021] In a further technical solution of the present invention, the cellulase activity in the enzymatic hydrolysis-assisted extraction step is 10000U / g-15000U / g, and the stirring interval during the enzymatic hydrolysis process is 30 min. Preferably, the ultrasonic power in the ultrasonic-enhanced extraction step is 400W, the frequency is 40kHz, and the extraction time is 40 min. Furthermore, in the decoction extraction step, the distilled water added during the first decoction is 7 times the weight of the mixed medicinal powder, and the distilled water added during the second decoction is 5 times the weight of the mixed medicinal powder, with both decoction times lasting 1.5 h.

[0022] In the alcohol precipitation purification step, the ethanol volume content of the purified alcohol is 60%, the standing temperature is 4℃-8℃, and the standing time is 24h. In the vacuum concentration step, the vacuum degree of the vacuum concentration is -0.08MPa to -0.09MPa, the temperature is 55℃-65℃, and the concentration is carried out to a relative density of 1.30-1.35, which is measured at 60℃. In the honey refining and paste collection step, the honey refining temperature is 110℃-115℃, the refining time is 5min-8min, the paste collection temperature is 80℃-85℃, and the criterion for the water droplet formation method is that the paste can be dropped into cold water and solidify into a bead that does not disperse.

[0023] In the quality inspection steps, the determination of the content of the indicator components includes: the content of Morinda officinalis oligosaccharide is not less than 1.5 mg / g, the content of Homalomena occulta volatile oil is not less than 0.8 mL / 100g, the content of Alpinia oxyphylla total flavonoids is not less than 3.0 mg / g, the content of Achyranthes bidentata polysaccharide is not less than 8.0 mg / g, the moisture content of the finished product is not more than 15%, and the total number of colonies in the microbial limit is not more than 1000 cfu / g, and the total number of molds and yeasts is not more than 100 cfu / g.

[0024] The beneficial effects of this invention are reflected in several aspects.

[0025] From the perspective of formulation composition, this invention selects *Homalomena occulta* and *Morinda officinalis* as principal herbs, with relatively large and comparable dosages of both, working together to exert the core effects of warming and tonifying kidney yang and strengthening tendons and bones. *Homalomena occulta* contains abundant volatile oils and diterpenoids, possessing the effects of dispelling wind and dampness, strengthening tendons and bones, and tonifying the liver and kidneys. Modern pharmacological studies have confirmed its anti-inflammatory, analgesic, and bone-forming effects. After being processed with brine, *Morinda officinalis* has an increased content of oligosaccharide components, enhancing its kidney-warming and yang-tonifying effects while reducing its drying properties. Studies have shown that *Morinda officinalis* oligosaccharides can significantly increase serum testosterone levels and improve sexual function. *Alpinia oxyphylla*, *Achyranthes bidentata*, *Euryale ferox*, and *Dioscorea opposita* are combined as assistant herbs. *Alpinia oxyphylla* warms the spleen and kidneys, consolidates essence and reduces urination; *Achyranthes bidentata* tonifies deficiency and moistens the lungs, strengthens tendons and invigorates collaterals; *Euryale ferox* and *Dioscorea opposita* together have the effects of strengthening the spleen and kidneys, consolidating essence and stopping diarrhea, working together to nourish the innate constitution through the strengthening of the acquired constitution. Achyranthes bidentata and cinnamon are used as adjuvant herbs. Achyranthes bidentata enters the liver and kidney meridians, tonifying the liver and kidneys, strengthening tendons and bones, guiding blood downwards, and guiding other herbs to the kidney meridian. Cinnamon warms and tonifies kidney yang, guiding fire back to its source. Citrus reticulata peel is used as the guiding herb, regulating qi and strengthening the spleen, harmonizing other herbs, and preventing cloying flavors from affecting the stomach. The entire formula embodies the characteristics of warming yang without causing dryness, tonifying without causing stagnation, and nourishing without being cloying, making it particularly suitable for the treatment of male menopausal syndrome and late-onset hypogonadism in the humid and hot climate of the Lingnan region.

[0026] From a process perspective, this invention employs a composite extraction technology combining enzymatic-assisted extraction, ultrasonic-enhanced extraction, and decoction extraction, significantly improving the extraction efficiency of active ingredients. Enzymatic hydrolysis disrupts the cell wall structure of medicinal materials, promoting the release of active ingredients. This is particularly effective for polysaccharide-rich herbs such as *Smilax glabra* and *Dioscorea opposita*, where enzymatic hydrolysis significantly increases polysaccharide yield. The cavitation effect of ultrasound further breaks down cell structures, accelerating the dissolution of active ingredients and shortening extraction time. Dual filtration clarification and alcohol precipitation purification effectively remove large molecular impurities such as starch and protein, improving product purity and stability. Vacuum concentration allows for rapid concentration at low temperatures, avoiding the damage of heat-sensitive components caused by high temperatures. Compared to traditional open-flame decoction, it better preserves volatile oils, flavonoids, and other active ingredients. The entire preparation process is scientifically sound, ensuring both the full extraction of active ingredients and maximum protection of heat-sensitive components, guaranteeing stable and controllable product quality.

[0027] From a quality control perspective, this invention establishes a quantitative control system for multiple indicators of components, including Morinda officinalis oligosaccharides, Homalomena occulta volatile oil, Alpinia oxyphylla total flavonoids, and Smilax glabra polysaccharides. Compared to existing technologies that rely solely on traditional morphological testing, this system offers more scientific, standardized, and controllable quality control. Quantitative control of multiple indicators comprehensively reflects product quality, ensuring consistent quality across batches.

[0028] From a pharmacological perspective, the ointment described in this invention exerts its kidney-tonifying, yang-boosting, and bone-strengthening effects through multiple targets and pathways. Morinda officinalis oligosaccharides can upregulate the expression of hypothalamic gonadotropin-releasing hormone, promote pituitary secretion of luteinizing hormone and follicle-stimulating hormone, thereby stimulating testicular interstitial cells to synthesize and secrete testosterone, increasing serum testosterone levels, and fundamentally improving gonadal function. Homalomena occulta volatile oil and cinnamaldehyde can promote relaxation of penile cavernous body smooth muscle, increase local blood flow, and improve erectile function. Alpinia oxyphylla extract can inhibit phosphodiesterase 5 activity and enhance erectile hardness. Achyranthes bidentata polysaccharides and Morinda officinalis oligosaccharides can promote osteoblast proliferation and differentiation, inhibit osteoclast activity, improve bone metabolism balance, and increase bone density. Achyranthes bidentata polysaccharides can increase muscle glycogen reserves, delay lactic acid accumulation, enhance mitochondrial oxidative phosphorylation function, increase ATP production efficiency, and exert an anti-fatigue effect. The combination of Euryale ferox and Alpinia oxyphylla can improve sperm density, motility, and the rate of normal morphology, thus improving semen quality.

[0029] From a clinical application perspective, the ointment described in this invention is specifically targeted at male menopausal syndrome and late-onset hypogonadism, with a clear therapeutic focus, unlike existing technologies that have a vague focus on urinary frequency, thus exhibiting stronger clinical specificity. The ointment is convenient to take, suitable for long-term use, and is suitable for middle-aged and elderly male patients.

[0030] From the perspective of regional characteristics, this invention makes full use of the authentic medicinal resources of Lingnan. Among them, salt-bajitian and Guangchenpi are the first batch of protected authentic medicinal materials of Lingnan, while Niudali, Rougui, Qianshi, and Yizhiren are the second batch of protected medicinal materials. It meets the characteristics of the hot and humid climate of Lingnan and the needs of men's health conditioning, and has clear regional medical characteristics and market competitive advantages.

[0031] In summary, the kidney-tonifying, muscle-strengthening, and yang-boosting formula provided by this invention has a reasonable formula, scientific process, controllable quality, and definite curative effect. It has significant advantages and broad application prospects in the prevention and treatment of male menopausal syndrome and late-onset hypogonadism. Detailed Implementation

[0032] The present invention will be further described in detail below with reference to specific embodiments. It should be noted that the following embodiments are for illustrative purposes only and are not intended to limit the scope of protection of the present invention. Unless otherwise specified, specific conditions in the embodiments are performed under conventional conditions or conditions recommended by the manufacturer. Reagents or instruments used, unless otherwise specified, are all commercially available conventional products.

[0033] All active pharmaceutical ingredients used in the examples conformed to the standards of the 2020 edition of the Chinese Pharmacopoeia. *Qiannianjian* was purchased from the Guangzhou Medicinal Herb Market; *Salted Morinda officinalis* was purchased from the Huazhou Medicinal Herb Market in Guangdong; *Alpinia oxyphylla* was purchased from the Dongfang Medicinal Herb Market in Hainan; *Achyranthes bidentata* was purchased from the Deqing County Medicinal Herb Market in Guangdong; *Euryale ferox* was purchased from the Suzhou Medicinal Herb Market in Jiangsu; *Dioscorea opposita* was purchased from the Huaiji County Medicinal Herb Market in Guangdong; *Achyranthes bidentata* was purchased from the Mianyang Medicinal Herb Market in Sichuan; *Cinnamomum cassia* was purchased from the authentic medicinal herb base in Guilin, Guangxi; *Citrus reticulata* was purchased from the Xinhui District Medicinal Herb Market in Guangdong; the honey was mature honey purchased from a bee farm in Conghua, Guangdong, with a moisture content of less than 20%; cellulase was purchased from Novozymes China Biotechnology Co., Ltd., with an enzyme activity of 15000 U / g; and ethanol was analytical grade and purchased from Sinopharm Chemical Reagent Co., Ltd.

[0034] Example 1

[0035] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula, consisting of 60g of *Homalomena occulta*, 60g of salt-processed *Morinda officinalis*, 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, 30g of *Achyranthes bidentata*, 15g of cinnamon, 15g of dried tangerine peel, and 450g of honey, which are boiled into a paste weighing 1000g. This formula is an optimal one, with a reasonable combination of principal, assistant, and adjuvant herbs, and the dosage of each herb is within an appropriate range, ensuring both the core efficacy of warming and tonifying kidney yang and avoiding excessive warming and drying.

[0036] The preparation method includes the following steps:

[0037] Step 1: Raw Material Pretreatment. The following nine medicinal herbs (60g of *Homalomena occulta*, 60g of *Morinda officinalis*, 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, 30g of *Achyranthes bidentata*, 15g of cinnamon, and 15g of dried tangerine peel) were subjected to quality inspection to ensure they met the standards of the Chinese Pharmacopoeia. Each herb was then pulverized separately using a high-speed pulverizer and passed through an 80-mesh sieve to obtain fine powder. *Homalomena occulta*, cinnamon, and dried tangerine peel, which contain volatile oils, were pulverized at low temperature. The pulverizer was frozen to 5°C during the pulverization process. Low-temperature pulverization effectively prevents the loss of volatile oils due to frictional heat during pulverization, ensuring the retention of volatile oil components. The pulverized herbs were then mixed evenly to obtain 360g of mixed herb powder.

[0038] Step 2, Enzymatic-Assisted Extraction. Take 360g of the above mixed herbal powder and add 2500mL of distilled water. The ratio of herbal powder to water is approximately 1:7. Stir thoroughly to ensure the herbal powder is evenly dispersed in the water. Add 1.08g of cellulase, which is 0.3% of the herbal weight. The activity of the cellulase used is 15000U / g. Adjust the pH to 4.8 with dilute hydrochloric acid or sodium hydroxide solution. This pH is the optimal pH for cellulase activity. Place the mixture in a constant temperature water bath at 45℃ and begin the enzymatic hydrolysis. Stir with a glass rod every 30 minutes during the hydrolysis process to ensure sufficient contact between the enzyme and the herbal material, thereby improving the hydrolysis efficiency. The total hydrolysis time is 1.5 hours. Enzymatic hydrolysis can disrupt the cellulose structure in the cell walls of the herbal materials, increasing cell wall permeability and promoting the release of active ingredients. Especially for herbs rich in polysaccharides, such as *Achyranthes bidentata* and *Dioscorea opposita*, enzymatic hydrolysis can significantly increase the polysaccharide yield.

[0039] Step 3: Ultrasonic Enhanced Extraction. After enzymatic hydrolysis, the hydrolysate is transferred to an ultrasonic extraction device and ultrasonically treated using an ultrasonic cleaner. The ultrasonic power is set to 400W and the frequency to 40kHz, which are commonly used parameters for ultrasonic extraction and produce a good cavitation effect. The ultrasonic extraction time is 40 minutes. The cavitation effect of ultrasound can further break down cell structures, generate microjets and shock waves, accelerate the diffusion of active ingredients from the cells into the solvent, shorten the extraction time, and improve the extraction efficiency. Temperature should be carefully controlled during ultrasonic treatment to avoid excessively high temperatures that could lead to the loss of heat-sensitive components.

[0040] Step 4: Decoction and Extraction. Transfer the ultrasonically treated extract to a stainless steel extraction vessel and heat to boiling. Initial heating can be done over high heat, but once boiling, immediately reduce to a simmer, maintaining a gentle boil between 95℃ and 100℃. Simmering over low heat avoids excessive heat that could destroy the active ingredients and ensures thorough extraction. During decoction, observe the liquid level frequently, adding any evaporated water to maintain a stable level and ensure the herbs are always submerged. The first decoction should last 1.5 hours. After decoction, filter the liquid through four layers of gauze and collect the filtrate. Do not discard the dregs; add 1800 mL of distilled water (approximately five times the weight of the dregs) and repeat the second decoction process as described above, also for 1.5 hours. The second decoction further extracts the remaining active ingredients from the dregs, increasing the yield. Filter and collect the filtrate from the second decoction. Combine the two filtrates to obtain approximately 3800 mL of liquid.

[0041] Step 5: Double Filtration Clarification. Allow the combined drug solution to cool to room temperature, which takes approximately 2-3 hours. After cooling, refrigerate the solution at 4°C for 12 hours. Low-temperature refrigeration causes some insoluble impurities, such as starch and protein, to precipitate. After removal, perform preliminary filtration using eight layers of gauze to remove coarse particles and precipitates. Then, perform fine filtration using a 0.45μm microporous membrane. The microporous membrane traps fine insoluble impurities and microorganisms, resulting in a clear and transparent drug solution. Double filtration effectively improves the purity of the drug solution, creating favorable conditions for subsequent alcohol precipitation and concentration.

[0042] Step Six: Alcohol Precipitation Purification. Slowly add 95% ethanol to the clarified drug solution while stirring continuously to avoid excessively high local ethanol concentrations that could lead to precipitation and loss of active ingredients. Accurately measure the ethanol volume using a graduated cylinder or pipette, and calculate the required amount of ethanol based on the drug solution volume to ensure a final ethanol volume content of 60%. For example, if the drug solution volume is 3800 mL, approximately 5700 mL of 95% ethanol should be added, resulting in a total volume of approximately 9500 mL and an ethanol volume content of 60%. After thorough mixing, place the precipitate in a 4°C refrigerator and let it stand for 24 hours. During this standing period, large molecules such as polysaccharides and proteins will gradually precipitate due to decreased solubility. After 24 hours of standing, filter through eight layers of gauze, discard the precipitate, and collect the clear filtrate. Alcohol precipitation purification effectively removes large molecular impurities, improving product purity and stability.

[0043] Step 7, Vacuum Concentration. Transfer the alcohol precipitate filtrate to a rotary evaporator, set the vacuum to -0.08 MPa and the water bath temperature to 60°C, and begin concentration. Under these vacuum and temperature conditions, the boiling point of ethanol is approximately 40°C-45°C, allowing for rapid evaporation and recovery. During concentration, continuously rotate the evaporator flask to form a thin film on the flask wall, increasing the evaporation area and improving concentration efficiency. First, recover ethanol until no alcohol odor remains, confirming that ethanol has been largely removed through olfactory judgment and refractive index testing. The recovered ethanol can be recycled, reflecting the concept of environmental protection. After ethanol recovery, continue concentrating the aqueous phase, adjusting the water bath temperature to 60°C and maintaining the vacuum at -0.08 MPa, until a thick paste is formed. Periodically sample and measure the relative density; stop concentration when the relative density reaches 1.32. The relative density is measured using the hydrostatic bottle method at 60°C. Vacuum concentration allows for rapid concentration at low temperatures, avoiding the damage of heat-sensitive components such as volatile oils and flavonoids caused by high temperatures, offering significant advantages over traditional open-flame cooking.

[0044] Step 8: Refining the Honey into a Thick Paste. Place 450g of high-quality honey in a stainless steel pot and heat over medium heat. As the temperature rises, the water in the honey begins to evaporate, and a large amount of foam will appear on the surface. When the temperature reaches 110℃, the foam gradually decreases, and the honey color deepens. At this point, maintain the temperature between 110℃ and 115℃ and continue heating for 5-8 minutes. Stir constantly during the honey refining process to prevent localized overheating and scorching. The refined honey will be dark brown, thick, and without any watery feel. Refining the honey removes excess water and impurities, allowing it to better blend with the medicinal paste. Cool the refined honey to about 80℃ and add the above-mentioned concentrated paste, stirring thoroughly until well mixed. Continue heating the mixture at 80℃-85℃ to thicken the paste, stirring constantly to prevent scorching and sticking to the pot. During the thickening process, periodically check the consistency using the water droplet test. Dip a small amount of paste into cold water with a glass rod and observe whether the paste can solidify into a bead that does not disperse. When the paste is dropped into water, it immediately solidifies into a round bead and retains its shape without spreading out in the water, indicating that the consistency is suitable. Continue reducing the paste until the total weight reaches 1000g, then stop heating.

[0045] Step Nine, Cooling and Filling. After the paste is collected, fill it quickly while it is still hot. At this point, the paste temperature is approximately 60℃-70℃, and its fluidity is good, making it easy to fill. The filling operation should be carried out in a clean environment. Before filling, sterilize the work surface with a UV lamp for 30 minutes. The filling containers are glass bottles with capacities of 250g and 500g, which have been sterilized at high temperature before use. Use a funnel or filling machine to fill the paste into the containers, accurately weighing each bottle to ensure consistent weight. Immediately after filling, tighten the cap to seal the bottle and invert it. The heat of the paste itself will create negative pressure, enhancing the sealing effect. Cool the bottle inverted to room temperature for approximately 6-8 hours. Do not move the bottle during the cooling process to avoid affecting the sealing effect. After cooling, turn the bottle upright and check the seal to ensure there is no leakage.

[0046] Step 10, Quality Inspection. The finished product must undergo comprehensive quality inspection to ensure it meets quality standards. Physical inspection: Observe the color, odor, and consistency of the paste. A qualified product should be brownish-red or dark brown, with a characteristic medicinal aroma, a fine and uniform texture, and no charring or off-odors. Moisture content determination: Use the drying method. Take 2g-3g of sample and dry it in an oven at 105℃ until constant weight. Calculate the moisture content. The moisture content of a qualified product should not exceed 15%. Microbiological limit testing: This includes total bacterial count, mold and yeast count, coliform bacteria, and pathogenic bacteria detection, conducted according to the methods of the 2020 edition of the Chinese Pharmacopoeia. The total bacterial count should not exceed 1000 cfu / g, and the total mold and yeast count should not exceed 100 cfu / g. Coliform bacteria, Salmonella, and Staphylococcus aureus should not be detected. Heavy metal and harmful element detection: Use atomic absorption spectrometry to detect the content of lead, cadmium, arsenic, mercury, and copper, which should comply with the standards of the Chinese Pharmacopoeia. The content of the indicator components was determined by high performance liquid chromatography (HPLC) to determine the oligosaccharide content of Morinda officinalis, steam distillation to determine the volatile oil content of Homalomena occulta, ultraviolet spectrophotometry to determine the total flavonoid content of Alpinia oxyphylla, and phenol-sulfuric acid method to determine the polysaccharide content of Smilax glabra. The ointment prepared in this embodiment was tested and found to contain 2.1 mg / g of Morinda officinalis oligosaccharide, 1.2 mL / 100g of Homalomena occulta volatile oil, 4.5 mg / g of Alpinia oxyphylla total flavonoids, 10.8 mg / g of Smilax glabra polysaccharide, and 12% moisture. Microbial limits were met, and the contents of heavy metals and harmful elements were all within the limits. All indicators met the quality standards.

[0047] Qualified finished products shall be labeled with information including product name, ingredients, indications, usage and dosage, precautions, production date, batch number, expiration date, and manufacturer. The outer packaging shall be a cardboard box, accompanied by a product instruction manual. Finished products should be stored in a cool, dry place, at a temperature not exceeding 20℃ and a relative humidity not exceeding 70%, with a shelf life of 24 months.

[0048] Example 2

[0049] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula, consisting of 50g of *Homalomena occulta*, 50g of salt-processed *Morinda officinalis*, 40g of *Alpinia oxyphylla*, 40g of *Achyranthes bidentata*, 30g of *Euryale ferox*, 30g of *Dioscorea opposita*, 20g of *Achyranthes bidentata*, 10g of cinnamon, 10g of dried tangerine peel, and 400g of honey, which are decocted into a 1000g paste. The dosages of each herb in this formula are taken at the lower limit of their respective ranges, making it suitable for patients with milder symptoms and reflecting the concept of individualized treatment.

[0050] The preparation method is basically the same as in Example 1, except that: in step one, 290g of mixed medicinal powder is taken; in step two, 2000mL of distilled water is added, the ratio of medicinal powder to water is about 1:7, and 0.87g of cellulase is added; in step four, 2000mL of distilled water is added for the first decoction, and 1450mL of distilled water is added for the second decoction, and the two filtrates are combined to obtain about 3200mL of medicinal liquid; in step six, about 4800mL of 95% ethanol is added to make the ethanol volume content reach 60%; in step seven, it is concentrated to a relative density of 1.30; in step eight, 400g of honey is taken and refined, and the paste is reduced to a total weight of 1000g.

[0051] The ointment prepared in this embodiment was tested and found to contain 1.6 mg / g of Morinda officinalis oligosaccharide, 0.9 mL / 100g of Homalomena occulta volatile oil, 3.2 mg / g of Alpinia oxyphylla total flavonoids, 8.5 mg / g of Smilax glabra polysaccharide, and 13% moisture. All other indicators met the quality standards. This embodiment verified the feasibility of the lower limit composition of the formula. Although the efficacy is relatively mild, the safety is higher, making it suitable for those with weak constitutions or first-time users.

[0052] Example 3

[0053] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula, consisting of 70g of *Homalomena occulta*, 70g of salt-processed *Morinda officinalis*, 60g of *Alpinia oxyphylla*, 60g of *Achyranthes bidentata*, 50g of *Euryale ferox*, 50g of *Dioscorea opposita*, 40g of *Achyranthes bidentata*, 20g of cinnamon, 20g of dried tangerine peel, and 500g of honey, which are decocted into a 1000g paste. The dosages of each herb in this formula are at the upper limit of their range, making it suitable for patients with more severe symptoms and stronger constitutions, providing a stronger warming and tonifying effect.

[0054] The preparation method is basically the same as in Example 1, except that: in step one, 430g of mixed medicinal powder is taken; in step two, 3000mL of distilled water and 1.29g of cellulase are added; in step four, 3000mL of distilled water is added for the first decoction and 2150mL of distilled water is added for the second decoction, and the two filtrates are combined to obtain about 4500mL of medicinal liquid; in step six, about 6750mL of 95% ethanol is added to make the ethanol volume content reach 60%; in step seven, it is concentrated to a relative density of 1.35; in step eight, 500g of honey is taken and refined at a temperature of 115℃ for 8 minutes, and the paste is reduced to a total weight of 1000g.

[0055] The ointment prepared in this embodiment was tested and found to contain 2.8 mg / g of Morinda officinalis oligosaccharide, 1.5 mL / 100g of Homalomena occulta volatile oil, 5.2 mg / g of Alpinia oxyphylla total flavonoids, 12.5 mg / g of Smilax glabra polysaccharide, and 14% moisture. All other indicators met the quality standards. This embodiment verifies the feasibility of the upper limit composition of the formula and has a stronger medicinal effect, but it has certain requirements for physical constitution. Those with yin deficiency and excessive fire should use it with caution.

[0056] Example 4

[0057] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula. The formula composition is the same as in Example 1, but the preparation method has been optimized in terms of enzymatic hydrolysis parameters. In step two, the cellulase activity is 10000 U / g, and the dosage is 0.3% of the medicinal material weight, i.e., 1.08g of cellulase is added. The pH is adjusted to 4.5, the hydrolysis temperature is 45℃, and the hydrolysis time is 1.5 hours. This embodiment verifies the hydrolysis effect under lower enzyme activity conditions. Although the enzyme activity is lower than in Example 1, a good extraction effect can still be achieved by appropriately extending the hydrolysis time, providing a reference for the selection of different enzyme sources. Other steps are the same as in Example 1.

[0058] The ointment prepared in this embodiment was tested and found to contain 1.9 mg / g of Morinda officinalis oligosaccharide, 1.1 mL / 100g of Homalomena occulta volatile oil, 4.2 mg / g of Alpinia oxyphylla total flavonoids, and 10.2 mg / g of Smilax glabra polysaccharide. All indicators met the quality standards and were similar to those in Example 1, demonstrating the stability and adaptability of the process.

[0059] Example 5

[0060] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula with the same composition as in Example 1, but the preparation method has been optimized in terms of concentration parameters. In step seven, the vacuum concentration is performed at a vacuum degree of -0.09 MPa and a temperature of 55°C, concentrating to a relative density of 1.30. This embodiment verifies the concentration effect under higher vacuum and lower temperature conditions. Under these conditions, ethanol recovery is faster and heat-sensitive components are better protected, but the concentration time may be slightly longer. Other steps are the same as in Example 1.

[0061] The ointment prepared in this embodiment was tested and found to contain 2.0 mg / g of Morinda officinalis oligosaccharide, 1.3 mL / 100g of Homalomena occulta volatile oil, 4.6 mg / g of Alpinia oxyphylla total flavonoids, and 10.5 mg / g of Smilax glabra polysaccharide. All indicators met the quality standards. It is worth noting that the Homalomena occulta volatile oil content in this embodiment is slightly higher than in Example 1, indicating that a lower concentration temperature is indeed beneficial for the retention of volatile oils.

[0062] Example 6

[0063] This embodiment prepares a kidney-tonifying, muscle-strengthening, and yang-boosting formula. The formula consists of 55g of *Homalomena occulta*, 65g of *Morinda officinalis* (salted), 45g of *Alpinia oxyphylla*, 55g of *Achyranthes bidentata*, 35g of *Euryale ferox*, 45g of *Dioscorea opposita*, 25g of *Achyranthes bidentata*, 12g of cinnamon, 18g of *Citrus reticulata*, and 475g of honey, which are boiled into a paste of 1000g. The dosage of each herb in this formula represents an intermediate value, further verifying the flexibility and adaptability of the formula. The preparation method is the same as in Example 1, with the material and solvent dosages adjusted accordingly based on the dosage of the raw materials. The paste prepared in this embodiment was tested, and all indicators met the quality standards. The oligosaccharide content of *Morinda officinalis* was 2.2mg / g, the volatile oil content of *Homalomena occulta* was 1.1mL / 100g, the total flavonoid content of *Alpinia oxyphylla* was 4.3mg / g, and the polysaccharide content of *Achyranthes bidentata* was 11.0mg / g, indicating stable and controllable quality.

[0064] Comparative Example 1

[0065] This comparative example prepares a paste with the following formula: 60g of salt-processed Morinda officinalis, 50g of Alpinia oxyphylla, 50g of Achyranthes bidentata, 40g of Euryale ferox, 40g of Dioscorea opposita, 30g of Achyranthes bidentata, 15g of Cinnamomum cassia, 15g of Citrus reticulata peel, and 450g of honey, which are boiled down to 1000g of paste. Compared with Example 1, this comparative example lacks one of the principal herbs, Homalomena occulta, reducing the total amount of medicinal materials by 60g. The amount of honey is correspondingly increased to 450g to maintain the total weight. The preparation method is the same as in Example 1, except that in step one, only Cinnamomum cassia and Citrus reticulata peel are pulverized at low temperature, as Homalomena occulta is absent.

[0066] The purpose of this comparative study is to verify the importance of *Homalomena occulta* in the formula. *Homalomena occulta* is rich in volatile oils and diterpenoids, possessing the effects of dispelling wind and dampness, strengthening muscles and bones, and tonifying the liver and kidneys. In the formula, it works together with salt-processed *Morinda officinalis* as the principal herbs, playing a core role in warming and tonifying kidney yang and strengthening muscles and bones. Without *Homalomena occulta*, the formula's warming and strengthening effects will be affected.

[0067] Comparative Example 2

[0068] This comparative example prepares an ointment with the following formula: 60g of *Homalomena occulta*, 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, 30g of *Achyranthes bidentata*, 15g of cinnamon, 15g of dried tangerine peel, and 450g of honey, which are boiled down to 1000g of ointment. Compared with Example 1, this comparative example lacks the other principal ingredient, *Morinda officinalis* (salted), but the other components are the same. The preparation method is the same as in Example 1.

[0069] The purpose of this comparative study is to verify the importance of salt-processed Morinda officinalis in the formulation. Salt-processed Morinda officinalis is one of the core medicinal materials of this invention; its oligosaccharide components can significantly increase serum testosterone levels and improve sexual function, making it a key medicinal material for treating male menopausal syndrome and late-onset hypogonadism. Without salt-processed Morinda officinalis, the formulation's regulatory effect on gonadal function will be significantly weakened.

[0070] Comparative Example 3

[0071] This comparative example prepares a paste with the following formula: 60g of *Homalomena occulta*, 60g of *Morinda officinalis* (salted), 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, and 450g of honey, which are boiled down to 1000g of paste. Compared with Example 1, this comparative example lacks the adjuvant herbs *Achyranthes bidentata*, cinnamon, and the guiding herb *Citrus reticulata*, while the other components are the same. The preparation method is the same as in Example 1, except that only *Homalomena occulta* is used for low-temperature pulverization in step one, as cinnamon and *Citrus reticulata* are absent.

[0072] The purpose of this comparative study is to verify the synergistic effect of Achyranthes bidentata, Cinnamomum cassia, and Citrus reticulata peel in the formula. Achyranthes bidentata guides blood downwards and directs other herbs to the kidney meridian; Cinnamomum cassia guides fire back to its source and warms and tonifies kidney yang; Citrus reticulata peel regulates qi, strengthens the spleen, and harmonizes other herbs. The combination of these three herbs embodies the wisdom of guiding herbs to their respective meridians and preventing excessive stagnation. Without these three herbs, the overall coordination and efficacy of the formula will be affected.

[0073] Comparative Example 4

[0074] This comparative example uses a traditional preparation method, with the same formulation as Example 1, but the preparation method differs. In step one, low-temperature pulverization is omitted; all medicinal materials are pulverized at room temperature. Steps two and three omit enzymatic extraction and ultrasonic-enhanced extraction, proceeding directly to the decoction extraction in step four. In step four, the first decoction time is extended to 2.5 hours, and the second decoction time is extended to 2 hours to compensate for the absence of enzymatic and ultrasonic steps. Step seven uses traditional open-flame decoction for concentration, without vacuum concentration, at a heating temperature of approximately 100°C, concentrating to a thick paste. Other steps are the same as in Example 1.

[0075] The purpose of this comparative example is to verify the superiority of the preparation process of this invention. While traditional processes are simple, they involve high extraction temperatures and long extraction times, which can easily lead to the loss of heat-sensitive components such as volatile oils, and also result in low extraction efficiency. By comparing with Example 1, it can be demonstrated that the application of modern pharmaceutical technologies such as enzymatically assisted extraction, ultrasonically enhanced extraction, and vacuum concentration can significantly improve product quality.

[0076] To evaluate the efficacy of the kidney-tonifying, muscle-strengthening, and yang-boosting formula described in this invention, the following pharmacodynamic experiments were conducted.

[0077] Experiment 1: Effect on serum testosterone levels

[0078] Sixty 8-week-old male SD rats, weighing 200g-220g, were randomly divided into 6 groups of 10 rats each: normal control group, model control group, Example 1 treatment group, Example 2 treatment group, Comparative Example 1 treatment group, and Comparative Example 2 treatment group. Except for the normal control group, all other groups underwent subcutaneous injection of hydrocortisone to establish a kidney-yang deficiency model for 21 consecutive days at a dose of 25mg / kg body weight daily. Starting from day 8, the normal control and model control groups were administered an equal volume of distilled water by gavage, while the treatment groups were administered the corresponding ointment-water solution by gavage. Based on the human equivalent dose, the dosage was 1.5g / kg body weight daily for 28 consecutive days. After the last administration, rats were fasted for 12 hours but allowed free access to water. Blood was collected from the orbital sinus, serum was separated, and serum testosterone levels were detected using enzyme-linked immunosorbent assay (ELISA).

[0079] Experimental results showed that the serum testosterone level in the normal control group was 6.8 ng / mL, while the serum testosterone level in the model control group was significantly reduced to 2.1 ng / mL, indicating successful model establishment. The serum testosterone level in the Example 1 treatment group was 5.6 ng / mL, in the Example 2 treatment group it was 4.8 ng / mL, in the Comparative Example 1 treatment group it was 3.9 ng / mL, and in the Comparative Example 2 treatment group it was 3.5 ng / mL. The serum testosterone levels in the Example 1 and Example 2 treatment groups were significantly higher than those in the model control group and the comparative example group, indicating that the ointment described in this invention can effectively increase the serum testosterone level in rats with kidney-yang deficiency. The effects of Comparative Example 1 and Comparative Example 2 were significantly weaker than those of Example 1 and Example 2, demonstrating that *Homalomena occulta* and *Morinda officinalis* play a key role in increasing testosterone levels, and the combination of the two produces a significant synergistic effect.

[0080] Experiment 2: Effect on bone mineral density

[0081] Sixty 8-week-old male SD rats were randomly divided into 6 groups of 10 each. The grouping and administration methods were the same as in Experiment 1. After 28 days of administration, the rats were sacrificed, and the femurs were harvested. The femoral bone mineral density was measured using a dual-energy X-ray absorptiometry (DXA) system.

[0082] Experimental results showed that the femoral bone mineral density of the normal control group rats was 0.215 g / cm³. 2 The femoral bone mineral density of the model control group rats was significantly reduced, to 0.168 g / cm³. 2 This indicates that kidney yang deficiency can lead to decreased bone mineral density. In Example 1, the femoral bone mineral density of rats in the treatment group was 0.202 g / cm³. 2 In Example 2, the dosage group was 0.192 g / cm³. 2 The control group (Comparative Example 1) had a drug concentration of 0.180 g / cm³. 2 The control group (Comparative Example 3) had a drug concentration of 0.175 g / cm³. 2The bone mineral density (BMD) of the treatment groups in Examples 1 and 2 was significantly higher than that of the model control group and the comparative group, indicating that the ointment described in this invention can effectively improve the BMD of rats with kidney-yang deficiency. Comparative Example 3, lacking Achyranthes bidentata, Cinnamomum cassia, and Citrus reticulata peel, showed a significantly weaker effect on improving BMD compared to the Example groups, indicating that these three herbs play an important synergistic role in strengthening tendons and bones.

[0083] Experiment 3: Effects on Fatigue Resistance

[0084] Sixty 8-week-old male mice, weighing 18-22g, were randomly divided into 6 groups of 10 mice each, as in Experiment 1. The administration method was the same as in Experiment 1: daily gavage administration of the corresponding ointment solution at a dose of 2g / kg body weight for 28 consecutive days. One hour after the last administration, a weight-bearing swimming test was conducted. Lead weights equivalent to 5% of the mice's body weight were attached to their tails, and the mice were placed in a swimming pool at 25℃. The time from the start of swimming to exhaustion was recorded, with exhaustion defined as the head being submerged in water for more than 10 seconds.

[0085] The experimental results showed that the exhaustive swimming time of mice in the normal control group was 45 minutes, in the model control group it was 28 minutes, in the Example 1 treatment group it was 42 minutes, in the Example 3 treatment group it was 48 minutes, in the Comparative Example 1 treatment group it was 35 minutes, and in the Comparative Example 2 treatment group it was 32 minutes. The exhaustive swimming time in the Example 1 and Example 3 treatment groups was significantly longer than that in the model control group and the comparative example group, indicating that the ointment described in this invention can effectively improve anti-fatigue ability. The effect of Example 3 was better than that of Example 1, indicating that the anti-fatigue effect was more significant at the upper limit of the formulation dosage. The effect of the comparative example group was significantly weaker than that of the example groups, indicating that *Homalomena occulta* and *Morinda officinalis* play important roles in anti-fatigue.

[0086] Experiment 4: Comparison of Quality Indicators

[0087] The content of index components in the ointments prepared in Example 1 and Comparative Example 4 was determined, and the results are shown in Table 1.

[0088] Table 1 Comparison of quality indicators between Example 1 and Comparative Example 4

[0089] sample Morinda officinalis oligosaccharides (mg / g) Millennium Health Volatile Oil (mL / 100g) Total flavonoids from Alpinia oxyphylla (mg / g) *Smilax glabra* polysaccharide (mg / g) Example 1 2.1 1.2 4.5 10.8 Comparative Example 4 1.6 0.6 3.5 8.2

[0090] As can be seen from Table 1, Example 1 uses modern preparation technologies such as enzymatic extraction, ultrasonic enhanced extraction, and vacuum concentration. The content of each component is significantly higher than that of Comparative Example 4, which is prepared using traditional processes. In particular, the content of the volatile oil of *Hypericum perforatum* in Example 1 is twice that of Comparative Example 4. This shows that the preparation process of the present invention can significantly improve the extraction efficiency and retention rate of the effective components and ensure product quality.

[0091] The comprehensive experimental data are summarized in Table 2.

[0092] Table 2 Summary of pharmacodynamic experimental data for each example and comparative example

[0093] Group Serum testosterone (ng / mL) <![CDATA[Femoral bone density (g / cm 2 )]]> Swimming time to exhaustion (min) normal control group 6.8 0.215 45 Model control group 2.1 0.168 28 Example 1 5.6 0.202 42 Example 2 4.8 0.192 - Example 3 - - 48 Comparative Example 1 3.9 0.18 35 Comparative Example 2 3.5 - 32 Comparative Example 3 - 0.175 -

[0094] Note: "-" indicates that the group did not participate in this experiment.

[0095] As can be clearly seen from Table 2, the example group is significantly better than the comparative group in all pharmacodynamic indicators, proving the scientific nature and superiority of the formulation and preparation process described in this invention.

[0096] The pharmacodynamic mechanism of the kidney-tonifying, muscle-strengthening, and yang-boosting formula described in this invention is manifested at multiple levels. At the molecular level, the oligosaccharide components in *Morinda officinalis* can upregulate the gene expression of hypothalamic gonadotropin-releasing hormone, promoting the pituitary gland to secrete luteinizing hormone and follicle-stimulating hormone. These two hormones act on testicular interstitial cells, activating key enzymes such as cholesterol side-chain cleavage enzymes, and promoting the biosynthesis of testosterone from cholesterol. The volatile oil components in *Homalomena occulta* can inhibit aromatase activity, reducing the conversion of testosterone to estradiol, thereby increasing serum free testosterone levels. The diarylheptane compounds in *Alpinia oxyphylla* have a similar effect to phosphodiesterase 5 inhibitors, increasing the concentration of cyclic guanosine monophosphate in penile cavernous body smooth muscle cells, promoting smooth muscle relaxation, and improving erectile function.

[0097] At the cellular level, Achyranthes bidentata polysaccharides can activate the bone morphogenetic protein signaling pathway in osteoblasts, promoting osteoblast proliferation and differentiation, and upregulating the expression of osteogenic markers such as osteocalcin and alkaline phosphatase. Simultaneously, Morinda officinalis oligosaccharides can upregulate osteoprotegerin expression, downregulate nuclear factor κB receptor activator ligand expression, inhibit osteoclast formation and activity, and reduce bone resorption. This bidirectional regulation of osteogenic and osteoclast activity tilts the bone metabolism balance towards bone formation, thereby increasing bone density and improving osteoporosis.

[0098] From an energy metabolism perspective, *Achyranthes bidentata* polysaccharides can promote the uptake and utilization of glucose by muscle cells, increase muscle glycogen reserves, and provide a sufficient energy source for exercise. Simultaneously, *Achyranthes bidentata* polysaccharides can enhance mitochondrial oxidative phosphorylation function, improve ATP synthesis efficiency, and delay lactic acid accumulation during exercise, thus exerting an anti-fatigue effect. Cinnamaldehyde in cinnamon can activate adenosine monophosphate-activated protein kinase, promote fatty acid oxidation, and increase energy supply.

[0099] From the perspective of anti-oxidative stress, the flavonoids in Euryale ferox and Alpinia oxyphylla have strong antioxidant activity, which can scavenge free radicals, reduce the damage of oxidative stress to sperm, and improve sperm density, motility, and normal morphology rate. The polysaccharides in Dioscorea opposita can enhance the activity of antioxidant enzymes such as superoxide dismutase and catalase, protecting cells from oxidative damage.

[0100] From the perspective of Traditional Chinese Medicine (TCM) theory, the formula of this invention embodies the principle of monarch-minister-assistant-guide in drug formulation. *Homalomena occulta* and *Morinda officinalis* (salted) serve as the monarch herbs, warming and tonifying kidney yang, strengthening tendons and bones, addressing the root cause of kidney yang deficiency. *Alpinia oxyphylla*, *Achyranthes bidentata*, *Euryale ferox*, and *Dioscorea opposita* serve as assistant herbs, strengthening the spleen and kidneys, consolidating essence and reducing urination, promoting the balance of acquired and innate constitutions. *Achyranthes bidentata* and cinnamon serve as adjuvant herbs, guiding the medicine to its proper channels and enhancing its efficacy. *Citrus reticulata* (dried tangerine peel) serves as the guide herb, regulating qi, strengthening the spleen, harmonizing the other herbs, and preventing cloying or irritating effects on the stomach. The entire formula embodies the characteristics of warming yang without causing dryness, tonifying without causing stagnation, and nourishing without being cloying, making it particularly suitable for the treatment of male menopausal syndrome and late-onset hypogonadism in the humid and hot climate of the Lingnan region.

[0101] The preparation process of this invention also embodies several innovations. Enzymatic hydrolysis-assisted extraction utilizes cellulase to disrupt cell wall structure, promoting the release of active ingredients, especially for macromolecules such as polysaccharides, where enzymatic hydrolysis significantly improves yield. Ultrasonic-enhanced extraction utilizes the cavitation and mechanical effects of ultrasound to accelerate the diffusion of active ingredients from the cell into the solvent, shortening extraction time and improving efficiency. Dual filtration clarification and alcohol precipitation purification effectively remove macromolecular impurities such as starch and protein, improving product purity and stability. Vacuum concentration is carried out under low temperature and low pressure conditions, maximizing the protection of heat-sensitive components such as volatile oils and flavonoids, ensuring product quality. The entire preparation process is scientifically sound and the process parameters have been optimized, ensuring both the full extraction of active ingredients and the maximum protection of heat-sensitive components, achieving stable and controllable product quality.

[0102] The experimental data from the above examples and comparative examples demonstrate that the kidney-tonifying, muscle-strengthening, and yang-boosting formula of this invention is significantly superior to the comparative example in increasing serum testosterone levels, improving bone density, and enhancing anti-fatigue ability. This fully proves the scientific nature of the formula and the superiority of the preparation process of this invention. This invention fully utilizes the authentic medicinal resources of Lingnan, targeting the pathogenesis of male menopausal syndrome and delayed-onset hypogonadism. Prepared using modern pharmaceutical technology, it features a rational formula, scientific process, controllable quality, and definite efficacy, and has broad application prospects in the field of men's health.

[0103] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A formula for tonifying the kidneys, strengthening muscles and bones, and boosting yang, characterized in that... It is prepared from the following raw materials in parts by weight: 50g-70g of *Homalomena occulta*, 50g-70g of *Morinda officinalis* (salted), 40g-60g of *Alpinia oxyphylla*, 40g-60g of *Achyranthes bidentata*, 30g-50g of *Euryale ferox*, 30g-50g of *Dioscorea opposita*, 20g-40g of *Achyranthes bidentata*, 10g-20g of cinnamon, 10g-20g of dried tangerine peel, and 400g-500g of honey, which are boiled down to make 1000g of paste.

2. The kidney-tonifying, muscle-strengthening, and yang-boosting formula according to claim 1, characterized in that... The raw materials consist of the following components by weight: 60g of *Homalomena occulta*, 60g of *Morinda officinalis* (salted), 50g of *Alpinia oxyphylla*, 50g of *Achyranthes bidentata*, 40g of *Euryale ferox*, 40g of *Dioscorea opposita*, 30g of *Achyranthes bidentata*, 15g of cinnamon, 15g of dried tangerine peel, and 450g of honey, which are boiled down to make 1000g of ointment.

3. The preparation method of the kidney-tonifying, muscle-strengthening, and yang-boosting formula according to claim 1, characterized in that... Includes the following steps: Step 1, Raw material pretreatment: Weigh the nine medicinal materials, namely, Homalomena occulta, Morinda officinalis, Alpinia oxyphylla, Achyranthes bidentata, Euryale ferox, Dioscorea opposita, Achyranthes bidentata, Cinnamomum cassia, and Citrus reticulata, according to their weight proportions and then pulverize them separately through an 80-mesh sieve. Among them, Homalomena occulta, Cinnamomum cassia, and Citrus reticulata are pulverized at a low temperature of 5℃-10℃. Mix the pulverized medicinal materials evenly to obtain mixed medicinal powder. Step 2, Enzymatic hydrolysis-assisted extraction: Take 360g of the mixed medicinal powder, add 2500mL of distilled water, add cellulase with an enzyme activity of 10000U / g or higher, and use 0.3% of the weight of the medicinal material. Adjust the pH value to 4.5-5.0, and enzymatically hydrolyze in a constant temperature water bath at 45℃ for 1.5h, stirring once every 30min during the enzymatic hydrolysis process; Step 3, Ultrasonic Enhanced Extraction: Transfer the enzymatic hydrolysate obtained in Step 2 to an ultrasonic extraction device and extract ultrasonically for 40 minutes at an ultrasonic power of 400W and a frequency of 40kHz. Step 4, decoction and extraction: Heat the extract obtained in step 3 to boiling, then simmer over low heat at 95℃-100℃ for 1.5 hours, replenishing the evaporated water in time to keep the liquid level stable. After decoction, filter through four layers of gauze and collect the filtrate. Add 1800mL of distilled water to the dregs and decoct for a second time for 1.5 hours. Filter and collect the filtrate. Combine the two filtrates to obtain 3500mL-4000mL of medicinal liquid. Step 5, double filtration clarification: After cooling the drug solution obtained in step 4 to room temperature, place it in a refrigerator at 4℃-8℃ for 12 hours. After taking it out, first perform preliminary filtration with eight layers of gauze, and then perform fine filtration with a microporous membrane with a pore size of 0.45μm to obtain a clear drug solution. Step 6, alcohol precipitation purification: Slowly add 95% ethanol to the clear drug solution obtained in step 5 while stirring, adjust the ethanol volume content to 60%, let it stand at 4℃-8℃ for 24 hours, filter it with eight layers of gauze, discard the precipitate, and collect the filtrate. Step 7, Vacuum Concentration: The filtrate obtained in Step 6 is concentrated under a vacuum of -0.08MPa to -0.09MPa and a temperature of 55℃ to 65℃. Ethanol is first recovered until there is no alcohol odor, and then the concentration is continued until a thick paste with a relative density of 1.30 to 1.35 is obtained. The relative density is measured at 60℃. Step 8, Refining and thickening the honey: Take 450g of honey and heat it to 110℃-115℃ for 5min-8min to refine the honey. After cooling to 80℃, add the concentrated paste obtained in Step 7 and mix thoroughly. Continue heating and thickening the paste at 80℃-85℃ while stirring constantly. Use the droplet bead method to check the consistency. Thicken the paste to a total weight of 1000g. Step 9, Cooling and Filling: While the paste obtained in Step 8 is still hot, quickly fill it into sterilized containers at 60℃-70℃, with each bottle containing 250g or 500g. After filling, immediately seal the container and invert it to cool to room temperature. Step 10, Quality Inspection: The finished product undergoes appearance inspection, moisture determination, microbial limit testing, heavy metal and harmful element detection, and indicator component content determination. After passing the inspection, the finished product is packaged.

4. The preparation method according to claim 3, characterized in that... The cellulase activity described in step two is 10000 U / g-15000 U / g, and the stirring interval during the enzymatic hydrolysis process is 30 min.

5. The preparation method according to claim 3, characterized in that... The ultrasonic extraction in step three uses an ultrasonic power of 400W, a frequency of 40kHz, and an extraction time of 40min.

6. The preparation method according to claim 3, characterized in that... In step four, the distilled water added during the first decoction is 7 times the weight of the mixed medicinal powder, and the distilled water added during the second decoction is 5 times the weight of the mixed medicinal powder. The decoction time for both times is 1.5 hours.

7. The preparation method according to claim 3, characterized in that... In step six, the ethanol volume content of the alcohol precipitation purification is 60%, the standing temperature is 4℃-8℃, and the standing time is 24h.

8. The preparation method according to claim 3, characterized in that... In step seven, the vacuum concentration is performed at a vacuum level of -0.08 MPa to -0.09 MPa and a temperature of 55°C to 65°C, concentrating the material to a relative density of 1.30 to 1.35, wherein the relative density is measured at 60°C.

9. The preparation method according to claim 3, characterized in that... The honey refining temperature in step eight is 110℃-115℃, the honey refining time is 5min-8min, the paste-forming temperature is 80℃-85℃, and the criterion for the drop-water bead method is that the paste can be dropped into cold water and solidify into a bead without spreading out.

10. The preparation method according to claim 3, characterized in that... The determination of the content of the index components mentioned in step ten includes that the content of Morinda officinalis oligosaccharide is not less than 1.5 mg / g, the content of Homalomena occulta volatile oil is not less than 0.8 mL / 100g, the content of Alpinia oxyphylla total flavonoids is not less than 3.0 mg / g, and the content of Achyranthes bidentata polysaccharide is not less than 8.0 mg / g. The moisture content of the finished product is not more than 15%, and the total number of colonies in the microbial limit is not more than 1000 cfu / g, and the total number of molds and yeasts is not more than 100 cfu / g.