Application of beer yeast peptide in improvement of sarcopenia

Abstract

The invention relates to application of beer yeast peptide in improving sarcopenia caused by aging, and belongs to the field of food science and nutriology. The beer yeast peptide takes beer yeast as a raw material, is obtained through complex enzyme enzymolysis and purification processes, and has low molecular weight and good bioavailability. The beer yeast peptide improves the activity of antioxidant enzymes (SOD and GSH-Px) in muscular tissues and serum, reduces the level of lipid peroxidation products (MDA), and enhances the oxidation resistance; meanwhile, the expression of inflammatory factors IL-6 and TNF-alpha is inhibited, and the anti-inflammatory effect is achieved, so that the sarcopenia caused by aging is improved, and the muscle function and quality are improved. Animal experiments prove that the traditional Chinese medicine composition has remarkable effects in the aspects of improving exercise tolerance, enhancing muscle grip strength, increasing the content of type I collagen and the like. Theoretical basis and technical support are provided for development and application of the beer yeast peptide in the field of functional food.

Description

Technical Field

[0001] This invention relates to the fields of food science and nutrition, specifically to the application of a brewer's yeast peptide in improving muscle loss, and more particularly to its functional food use in regulating age-related sarcopenia through a dual mechanism of antioxidation and anti-inflammation, as well as its preparation method and detection technology. Background Technology

[0002] As my country's population ages, the prevention and control of age-related diseases has become an important social and public health issue. Sarcopenia, an age-related degenerative disease of skeletal muscle mass and function, is widely recognized as a significant factor affecting the quality of life and physical function of the elderly. Studies have shown that the occurrence of sarcopenia is closely related to decreased skeletal muscle protein synthesis capacity, increased oxidative stress, and chronic inflammation. Epidemiological data in China show that the prevalence of sarcopenia among people aged 60 and above ranges from 5.7% to 23.9%, and shows a gradually increasing trend with age.

[0003] In intervention strategies for sarcopenia, nutritional supplementation is considered one of the fundamental and crucial means. The Chinese Expert Consensus on Prevention and Intervention of Sarcopenia in the Elderly (2023) points out that malnutrition is a significant cause of decreased muscle protein synthesis, and the intake levels of protein, DHA, EPA, and vitamin D play a significant role in maintaining muscle mass in the elderly. However, due to the decline in digestive function in the elderly, their ability to digest and absorb whole protein decreases, making it difficult to fully utilize the essential amino acids in protein. Therefore, easily absorbed small molecule peptides become ideal nutritional intervention components.

[0004] Brewer's yeast peptides are low-molecular-weight peptides obtained from brewer's yeast through enzymatic hydrolysis and purification processes. They are not only rich in various essential amino acids but also possess excellent biological activity. Current research indicates that brewer's yeast protein has multiple physiological functions, including antioxidant activity, improved muscle mass, and reduced blood pressure. For example, population studies have shown that continuous supplementation with yeast protein powder for 8 weeks can significantly improve bench press ability, increase walking speed, and improve blood pressure levels, suggesting its potential application value in sports nutrition and muscle health.

[0005] Although existing studies have indicated that brewer's yeast proteins and peptides possess certain antioxidant capabilities and may exert their effects through mechanisms such as scavenging free radicals, inhibiting lipid oxidation, enhancing antioxidant enzyme activity, and activating the Keap1-Nrf2 pathway, systematic research on the function of brewer's yeast peptides in combating age-related sarcopenia is still lacking. In particular, their efficacy in alleviating sarcopenia symptoms under the synergistic regulation of both antioxidant and anti-inflammatory pathways remains unclear.

[0006] Furthermore, the lack of mechanistic validation of brewer's yeast peptides in improving muscle function, especially systematic review data in animal models, limits their development and application in the functional food field. Therefore, there is an urgent need to clarify the specific mechanism and effects of brewer's yeast peptides in improving sarcopenia through animal experimental evidence, providing a scientific basis for their application in food, nutrition, and health care. Summary of the Invention

[0007] To address the limitations of existing technologies in intervening in sarcopenia caused by oxidative stress and chronic inflammation, such as low nutrient absorption efficiency and a weak foundation for the application of functional foods, this invention provides a technical solution based on brewer's yeast peptides. This solution fully leverages the comprehensive physiological functions of brewer's yeast peptides in antioxidation and anti-inflammation, and, combined with animal experiments, verifies its effectiveness in improving muscle function, quality, and structure, thus providing a practical and feasible nutritional intervention for age-related sarcopenia. Based on this, the following describes in detail, with reference to several embodiments of this invention, the application methods, preparation methods, and evaluation criteria of brewer's yeast peptides in improving sarcopenia.

[0008] In one embodiment of the present invention, an application of brewer's yeast peptide in the preparation of a composition for improving sarcopenia is provided, wherein the brewer's yeast peptide works through the following dual mechanisms: (a) an antioxidant mechanism: enhancing antioxidant capacity by increasing the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in muscle tissue and serum and reducing malondialdehyde (MDA) levels; and (b) an anti-inflammatory mechanism: reducing chronic inflammation levels by inhibiting the expression of inflammatory factors interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α).

[0009] Furthermore, the brewer's yeast peptide can be prepared by the following process: (1) using brewer's yeast as raw material; (2) using a complex protease (including trypsin and flavor protease) to enzymatically hydrolyze for 4 hours at pH 7.0 and 50°C, and then pasteurizing at 95°C for 10 minutes after enzymatic hydrolysis to terminate the reaction; (3) obtaining small molecule peptides with a relative molecular mass of less than 5000 Da by membrane separation or other purification methods.

[0010] In one embodiment of the invention, the sarcopenia is age-related sarcopenia, and the composition is used to improve at least one of the following muscle function indicators: In the fatigue bar test, the bar time is extended by no less than 4 minutes (test conditions: rotation speed 20 rpm, initial height 5 cm). In the limb grip strength test, the grip strength maintenance time should be increased by no less than 1 minute (test conditions: 2 mm diameter metal wire, 30° tilt angle).

[0011] Furthermore, the composition can achieve muscle structure reconstruction and quality restoration by increasing the content of type I collagen in muscle tissue. Preferably, the integrated optical density (IOD) value of type I collagen in the tibialis anterior or gastrocnemius muscle is increased by more than 20%.

[0012] Preferably, the dosage of the brewer's yeast peptide is 50-200 mg / kg / day, and oral administration is more convenient, conforms to consumer usage habits, and is suitable for long-term nutritional intervention.

[0013] In one embodiment of the present invention, the brewer's yeast peptide can be used to construct a functional food for improving sarcopenia, the food comprising: The yeast peptide is used as the active ingredient; A pharmaceutically or food-acceptable carrier; The dosage form of the food can be selected as powder, tablet or liquid beverage to meet the needs of different consumption scenarios and groups.

[0014] Furthermore, to ensure the nutritional efficacy and quality stability of the functional food, the content of brewer's yeast peptides in the composition should account for 10%-90% of the total weight. Preferably, it should meet the following quality control standards: The total protein content shall not be less than 80% (Kjeldahl method). The antioxidant capacity EC50 value is not higher than 2 mg / mL (determined by DPPH free radical scavenging method).

[0015] In one embodiment of the present invention, a detection method for evaluating the improvement effect of sarcopenia is provided, comprising the following steps: Changes in the lean meat / fat ratio were detected using body composition analysis, with an increase in lean meat content of more than 5% considered an effective improvement. The changes in the content of type I and type III collagen in muscle tissue were detected by Sirius red staining. If the IOD value increased by more than 15%, it was considered effective.

[0016] In one embodiment of the present invention, a quality control method for brewer's yeast peptides is also proposed to ensure the stability of their composition and consistency of their biological activity, specifically including: Its molecular weight distribution was determined by liquid chromatography-mass spectrometry (LC-MS), with the main peak located in the range of 300-1000 Da; Its ability to activate the Nrf2 signaling pathway was detected by enzyme-linked immunosorbent assay (ELISA), and its activity was not less than 80% compared with the standard control.

[0017] Furthermore, the brewer's yeast peptide also has the function of significantly improving the motor ability of aged animals, manifested in the improvement of multiple motor performance indicators such as muscle wet weight ratio, grip strength duration, and rotarod time. At the same time, it can reduce oxidative stress levels and inflammatory response indicators such as IL-6, TNF-α, and MDA, thereby achieving multidimensional intervention on age-related muscle degenerative changes.

[0018] Based on the above technical solution, the application of the brewer's yeast peptide of the present invention in the preparation of compositions for improving sarcopenia effectively alleviates the problem of muscle mass decline and functional degeneration caused by aging through the synergistic effect of multiple targets. Specifically, the brewer's yeast peptide utilizes its dual antioxidant and anti-inflammatory mechanisms to intervene at the source of the main pathogenic factors of sarcopenia, namely oxidative stress and chronic inflammatory state.

[0019] In terms of antioxidant activity, brewer's yeast peptides can significantly enhance the activity of endogenous antioxidant enzymes such as SOD (superoxide dismutase) and GSH-Px (glutathione peroxidase) in the body, while reducing the content of lipid peroxidation products such as MDA (malondialdehyde), thereby enhancing the cell's ability to respond to oxidative stress and maintaining the structural and metabolic stability of muscle tissue. This mechanism helps to inhibit myofiber damage and apoptosis caused by excessive free radical production.

[0020] In terms of anti-inflammation, brewer's yeast peptides reduce systemic and local inflammatory responses by inhibiting the expression of pro-inflammatory factors such as IL-6 and TNF-α, breaking the vicious cycle of inflammation-muscle degeneration and providing a favorable environment for muscle regeneration and functional recovery. Animal experiments have shown that after intervention with brewer's yeast peptides, the expression levels of inflammatory factors in the gastrocnemius muscle tissue of mice decreased significantly, and the muscle morphology and structure were significantly improved.

[0021] Furthermore, rotarod fatigue tests and limb grip strength tests conducted in animal models demonstrated that brewer's yeast peptides effectively improve exercise endurance and muscle strength, with a dose-dependent effect. The high-dose intervention group showed a rotarod time extension of over 4 minutes and an increase in limb grip strength of over 1 minute, significantly superior to the model group and the low-dose group, indicating good potential for functional improvement.

[0022] Histological analysis showed that brewer's yeast peptides can promote the expression of type I collagen in skeletal muscle tissue, with an increase in integrated optical density (IOD) exceeding 20%, indicating a positive effect on muscle structure repair. This result is directly related to muscle mass recovery and helps enhance the mechanical support and functional performance of muscle tissue.

[0023] Regarding quality control, this invention further proposes a method for detecting the composition and functional activity of brewer's yeast peptides. LC-MS analysis is used to analyze their molecular weight distribution, ensuring the main peak is concentrated in the 300-1000 Da range; and ELISA is used to detect their activation ability of the Nrf2 signaling pathway, providing a standardized basis for product stability and bioefficacy.

[0024] In practical applications, the brewer's yeast peptide of the present invention can be widely used in various scenarios such as sarcopenia in the elderly, postoperative rehabilitation, and muscle degenerative changes caused by chronic diseases. It is especially suitable as a key active ingredient in functional foods and can be developed into dosage forms such as powders, tablets or beverages to meet the nutritional intervention needs of different populations.

[0025] In summary, the brewer's yeast peptide provided by this invention and its application in improving sarcopenia innovatively integrate antioxidant and anti-inflammatory mechanisms, have clear experimental verification, good biosafety and broad application prospects, are suitable for industrial promotion, and are expected to play an important role in the fields of elderly health and sports nutrition. Attached Figure Description

[0026] Figure 1 This is a statistical graph showing the effects of brewer's yeast peptides on body composition (lean meat content, fat ratio, etc.) in aging model mice, illustrating its effect on improving muscle quality.

[0027] Figure 2 Images and integrated optical density analysis results of the changes in type I and type III collagen content in mouse muscle tissue by brewer's yeast peptides are used to illustrate its promoting effect on muscle structure reconstruction.

[0028] Figure 3 The graph shows the changes in SOD, GSH-Px activity and MDA content in mouse serum and muscle tissue after intervention with brewer's yeast peptides, illustrating its antioxidant mechanism.

[0029] Figure 4 The diagram shows the regulatory effect of brewer's yeast peptide on the levels of inflammatory factors IL-6 and TNF-α in mouse serum, reflecting its anti-inflammatory activity. Detailed Implementation

[0030] The technical solution of the present invention will be further described below through specific embodiments, but the present invention should not be limited to the following embodiments. Unless otherwise stated, the raw materials, reagents, experimental equipment and conventional technical means used in the following embodiments are well known to those skilled in the art.

[0031] Example 1: Establishment of an animal model and design of intervention groups In one embodiment of the present invention: Kunming mice (SPF grade), male, 8 weeks old, with an initial weight of 40±4 g, were selected and provided by Beijing Vital River Laboratory Animal Technology Co., Ltd. Prior to the experiment, the mice underwent 7 days of acclimatization in a clean-grade animal room under the following conditions: temperature (22±2)℃, relative humidity (50±10)%, 12 h light / 12 h dark cycle, and free access to standard pelleted feed and water.

[0032] The experiment was divided into 6 groups, with 6 animals in each group. The specific groupings are as follows: Blank control group (no modeling, no intervention); Model group (modeling, no intervention); Vitamin C positive control group; Low-dose yeast peptide group (50 mg / kg); Medium-dose group of yeast peptides (100 mg / kg); High-dose yeast peptide group (200 mg / kg).

[0033] Except for the blank control group, the other five groups of mice were injected subcutaneously into the neck and back of their necks daily with D-galactose (Sigma, purity ≥99%) solution at a dose of 300 mg / kg·d. The solution was prepared with sterile physiological saline, and the injection volume was 0.2 mL / d, for 8 consecutive weeks, to induce an aging model. Mice in the blank control group were injected with an equal volume of sterile physiological saline as a control.

[0034] While D-galactose modeling was being established, mice in each group were administered the corresponding intervention via oral gavage daily, with a uniform gavage volume of 0.5 mL / day. The intervention details are as follows: Blank control group and model group: were given an equal volume of purified water (Milli-Q). Vitamin C positive control group: Administered 200 mg / kg of vitamin C (Sinopharm Group, pharmacopoeia grade); Low-dose yeast peptide group: Administered 50 mg / kg brewer's yeast peptide solution; Medium-dose group of yeast peptides: 100 mg / kg brewer's yeast peptide solution was administered; High-dose yeast peptide group: Administered 200 mg / kg brewer's yeast peptide solution.

[0035]

[0036] Example 2: Preparation method of brewer's yeast peptides The brewer's yeast peptide was prepared in the laboratory, and the preparation process is as follows: 1. Raw material source: Yeast sludge from the beer brewing process, which is removed by centrifugation and washing, and then dried and stored; 2. Enzymatic hydrolysis step: Dissolve the freeze-dried yeast powder in a 2% protein concentration solution, adjust the pH to 7.0, and add a mixed complex enzyme of trypsin (Sigma-Aldrich) and flavor protease (Novozymes) at a concentration of 1.5% (w / w). 3. Reaction conditions: Enzymatic hydrolysis was carried out in a water bath with shaking at 50℃ for 4 hours. After the enzymatic hydrolysis was completed, the reaction was terminated by pasteurization at 95℃ for 10 minutes. 4. Separation and purification: Ultrafiltration membrane separation technology was used to retain peptides with a molecular weight cutoff of 5 kDa. Peptides with a molecular weight of <5000 Da were collected, concentrated under reduced pressure, and then freeze-dried. 5. Testing indicators: Molecular weight distribution: Measured by liquid chromatography-mass spectrometry (LC-MS), the main peaks were concentrated in the range of 300-1000 Da; Protein content: Determined by the Kjeldahl method, total protein content ≥80%; Antioxidant capacity: Its antioxidant activity was determined using the DPPH free radical scavenging assay, and the result was EC50≤2 mg / mL; Bioactivity validation: Its Nrf2 pathway activation ability was assessed by ELISA, with an activity ≥80% compared to the standard control.

[0037] The yeast peptides were prepared as stable solutions before intervention and stored at 4°C in the dark to ensure that their components did not degrade during administration. Fresh solutions were prepared daily to ensure the stability of the active ingredients.

[0038] The entire intervention period lasted 8 weeks, during which subcutaneous injections and gavage were performed at fixed times daily (9:00 ± 0.5 h). Animals were weighed weekly during the intervention period, and their eating behavior, mental state, activity level, and coat condition were observed. Any adverse reactions or deaths were recorded.

[0039] This implementation plan is used to systematically verify the feasibility and dose-dependent nature of the effects of brewer's yeast peptides in improving sarcopenia, enhancing muscle function, and exhibiting antioxidant and anti-inflammatory effects in vivo. Subsequent assessments will utilize a comprehensive evaluation of its biological effects through functional testing, biochemical analysis, tissue staining, and image analysis.

[0040] Available options: During the intervention, the mice were monitored weekly for body weight and their mental state (activity, lethargy), food and water intake, and general physiological indicators such as fur luster were assessed to determine the model establishment and intervention effect. Mice with a body weight loss of more than 15%, abnormal behavior, or severe piloerection were immediately removed from the experimental group.

[0041] At the end of week 8 of the experiment, 24 hours after the last intervention, muscle function assessment and tissue sampling were performed, including the following two behavioral function tests.

[0042] Example 3: Behavioral Functional Evaluation in Mice Muscle function assessment (see Table 2) The motor ability and muscle function of mice were quantitatively evaluated using a rotarod fatigue tester and a grip strength meter.

[0043] (1) Fatigue rotating rod experiment: Experimental equipment: Rotary bar fatigue tester (ZB-200 model, Beijing Zhongyi Kexin); Parameter settings: Rotation speed is set to 20 rpm, rotor diameter is 3 cm, and height from the ground is 50 cm; Procedure: Place the mouse on the rotating bar, start timing, and record the time (in minutes) during which the mouse remains on the bar. Each animal should be tested only once. Maintain a quiet experimental environment to avoid interference during the test.

[0044] Evaluation criteria: If the mouse falls three times consecutively, the test will be terminated, and the time of the first fall will be taken as the final result.

[0045] Experimental results: The average time spent on the stick in the model group was significantly shorter than that in the blank control group (P<0.01), suggesting that the aging model leads to a decline in muscle endurance. The time spent in the yeast peptide intervention group was longer than that in the model group, and this trend was dose-dependent. The high-dose group had an average duration of 4.58 min longer, recovering to near-normal levels (P<0.01).

[0046] (2) Limb grip strength test: Experimental equipment: Limb grip strength tester (YLS-13A, Jinan Yizhong Biotechnology); Testing setup: Inclined plane angle set at 30°, metal wire diameter at 2 mm, surface smooth and non-slipping; Operating procedure: Gently place the mouse at the bottom of the test ramp. The mouse will naturally and instinctively climb the metal wire, and the timer will start. When the mouse's limbs detach from the metal wire, stop the timer and record the holding time. Each animal was tested 3 times, with 2 minutes between tests, and the average value was taken as the final result. Experimental results: The average gripping time in the model group was significantly shorter than that in the blank control group, indicating decreased muscle strength. After intervention with yeast peptides, especially in the high-dose group, the grip strength and time of mice increased significantly; The high-dose group showed an improvement of approximately 1.18 min compared to the model group (P<0.05), demonstrating a significant improvement in muscle function. The results were similar to those of the vitamin C group, indicating that yeast peptides have similar functional effects to classic antioxidants.

[0047]

[0048] As shown in Table 2, the fatigue rotarod time and grip strength maintenance time of the model group mice were significantly lower than those of the blank group, which verified the successful construction of the aging model of muscle function decline. After intervention with brewer's yeast peptide, the relevant indicators were significantly improved and showed a dose-dependent trend. The effect of the dose group was better than that of the vitamin C group.

[0049] The above behavioral tests are one of the key means of verifying the function of this invention. Combined with subsequent histological and molecular indicators, they can comprehensively reflect the overall efficacy of brewer's yeast peptides in improving sarcopenia.

[0050] Example 4: Mouse body composition detection Body composition analysis (see Figure 1 ) The ratio of lean meat to fat in mice after the intervention was quantitatively determined using a low-field nuclear magnetic resonance small animal body composition analyzer (NMI20-040H-I, small nuclear magnetic resonance, Shanghai Xintuo).

[0051] Detection method: Each mouse was fasted for 12 hours and then weighed. The animals were then placed individually in a special mouse testing chamber (red cage). Adjust the position of the fixing rod to naturally restrain the mouse's limb movements and ensure that there is no violent struggle during the measurement process; Push the mouse cage into the center of the instrument cavity and start scanning to obtain parameters such as lean body mass, fat weight and total water content; Each animal was measured once, and the data was automatically analyzed and output by the built-in software.

[0052] Analysis of experimental results: The body composition of the model group mice showed abnormal changes: fat content increased and lean meat content decreased slightly; After intervention with yeast peptides, all dosage groups showed a trend of increased lean meat content, especially the medium-dose and high-dose groups. Compared with the model group, the lean meat / fat ratio was significantly improved in the vitamin C group and the high-dose group, suggesting that yeast peptides have a positive effect on regulating muscle metabolism.

[0053] Example 5: Detection of collagen expression in muscle tissue Collagen testing (see) Figure 2 ) To evaluate the effect of brewer's yeast peptides on muscle tissue structure, the expression levels of type I and type III collagen in mouse gastrocnemius muscle were detected by Sirius red staining and immunohistochemistry (IHC) staining.

[0054] Staining steps and image analysis: 1. Paraffin section preparation: After the gastrocnemius muscle tissue was removed, it was fixed in 4% paraformaldehyde for 24 h, dehydrated, cleared and then embedded in paraffin. It was then cut into 10 μm thick continuous transverse sections using a Leica RM2235 microtome. 2. Sirius red staining: The Zhongshan Jinqiao reagent kit was used. After staining for 1 hour, the slides were dehydrated with anhydrous ethanol, cleared, and mounted. 3. Immunohistochemical staining: Type I collagen antibody (Wuhan Sanying, batch number: 66761-1-Ig) was used, and the procedure was performed according to the standard SP method; 4. Image acquisition and analysis: Images were taken using a Leica microscope and imported into Image-Pro Plus 6.0 software. The integrated optical density (IOD) values ​​of the red (type I) and yellow (type III) collagen signals in each image were measured.

[0055] Analysis of experimental results: The content of both type I and type III collagen in the muscle tissue of the model group was significantly reduced, suggesting that aging causes damage to collagen structure. The yeast peptide medium- and high-dose groups, as well as the vitamin C group, all showed a significant increase in the IOD value of type I collagen. Among them, the IOD value of type I collagen in the high-dose group increased by more than 20%, which was statistically significant (P<0.05). This indicates that brewer's yeast peptides help restore skeletal muscle matrix components and improve tissue structural stability.

[0056] Example 6: Antioxidant Capacity Test Antioxidant index detection (see) Figure 3 ) This invention further evaluates the ability of brewer's yeast peptides to regulate antioxidant state by detecting the activity of antioxidant-related enzymes and the level of lipid peroxides.

[0057] Test samples and methods: Mouse serum and gastrocnemius muscle tissue homogenates (mass ratio 1:9, PBS buffer) were collected and detected by colorimetric method using a commercial kit (Nanjing Jiancheng Biotechnology Co., Ltd.). The indicators tested include: SOD (superoxide dismutase) activity; GSH-Px (glutathione peroxidase) activity; MDA (malondialdehyde) content; The detection wavelengths were: SOD (550 nm), GSH-Px (412 nm), and MDA (532 nm). Each sample was tested in 3 replicate wells, and the average value was taken as the final result.

[0058] Analysis of experimental results: The activity of SOD and GSH-Px in the model group decreased significantly, while the level of MDA increased significantly (P<0.01), indicating enhanced oxidative stress. Yeast peptide intervention significantly improved the above indicators: SOD and GSH-Px activities were significantly increased in the medium and high dose groups; MDA levels decreased significantly in the medium and high dose groups, while in the high dose group they recovered to levels close to the blank control. The study suggests that brewer's yeast peptides can effectively enhance the antioxidant defense system and reduce the risk of oxidative damage.

[0059] Example 7: Detection of Inflammatory Factor Levels Inflammatory factor detection (see Figure 4 ) The expression levels of two key inflammatory factors, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), in mouse serum were detected by enzyme-linked immunosorbent assay (ELISA).

[0060] Operating procedures: The mouse IL-6 (batch number: Ml002293A) and TNF-α (batch number: Ml002095A) ELISA detection kits provided by Enzyme-Link Biotechnology Co., Ltd. were used. Serum samples were stored at -80°C and thawed thoroughly and centrifuged to remove impurities before testing. Add the standard, the sample to be tested and the enzyme conjugate to each well, and complete the incubation, washing and color development according to the instructions. Absorbance values ​​were read at a wavelength of 450 nm, and the concentrations of each factor were calculated.

[0061] Analysis of experimental results: The serum levels of IL-6 and TNF-α in the model group were significantly elevated, suggesting the presence of a chronic low-grade inflammatory response in the aging state; The medium and high doses of yeast peptides significantly reduced these two inflammatory markers, with effects comparable to those of vitamin C. In the high-dose group, the levels of IL-6 and TNF-α decreased by more than 25%, with significant differences (P<0.01). This indicates that brewer's yeast peptides alleviate chronic inflammation in tissues by regulating inflammatory pathways, thus contributing to muscle function recovery.

[0062] Example 8: Quantitative Analysis Method for Collagen in Muscle Tissue Preferred: In this embodiment of the invention, the gastrocnemius muscle was selected as the observation subject to verify the histological effect of brewer's yeast peptides in improving muscle structure. As a typical weight-bearing muscle group of the lower limbs, the gastrocnemius muscle is rich in type I and type II muscle fibers and is an important tissue for evaluating muscle structural integrity and fiber arrangement.

[0063] In the experiment, the gastrocnemius muscle tissue extracted after yeast peptide intervention was processed and analyzed as follows: 1. Fixation and embedding: Immediately after the muscle tissue was excised, it was placed in 4% paraformaldehyde solution and fixed at 4°C for 24 hours. After dehydration with graded ethanol and clearing with xylene, it was embedded in paraffin. 2. Section preparation: Using a Leica RM2235 paraffin microtome, cross-sections with a thickness of 5-7 μm were continuously cut; 3. Immunohistochemical staining: Type I collagen antibody (Wuhan Sanying, batch number: 66761-1-Ig) was used at a working concentration of 1:200. Following the SP kit procedure, antigen retrieval, blocking, primary antibody incubation, secondary antibody labeling, DAB staining, and hematoxylin counterstaining were performed. 4. Image Acquisition and Analysis: Five non-overlapping fields of view were photographed for each group of slices using a Leica DMi8 microscope. Import the image into Image-Pro Plus 6.0 image analysis software, set a uniform threshold and recognition parameters, and measure the integrated optical density (IOD) value of the brown-yellow region in each image. The average IOD value is used as a quantitative indicator of type I collagen content to reflect the degree of muscle tissue repair.

[0064] The experimental results showed that the IOD value of type I collagen in the gastrocnemius muscle of the high-dose brewer's yeast peptide intervention group increased by more than 20%, which was statistically significant compared with the model group (P<0.05). This indicates that the peptide helps promote muscle tissue reconstruction and improve the rupture of interstitial fibers and collagen loss caused by aging.

[0065] Example 9: Quality control method for brewer's yeast peptides Furthermore: To ensure the functional consistency, quality stability, and traceability of brewer's yeast peptide products during industrial application, this invention establishes the following quality control standard system: 1. Molecular weight distribution detection: A liquid chromatography-mass spectrometry (LC-MS) system equipped with a C18 reversed-phase column was used. The mobile phase consisted of a gradient elution of 0.1% formic acid aqueous solution and acetonitrile; The injection volume was 10 μL, and the detection wavelength was 214 nm. The results showed that the main peak of the target yeast peptide was concentrated in the 300~1000 Da range, accounting for more than 80% of the total area. Small molecule peptides within this range possess excellent water solubility, tissue permeability, and intestinal absorption efficiency, which are the basis for achieving functional activity.

[0066] 2. Bioactivity assay: The activation capacity of yeast peptides on the Keap1-Nrf2 antioxidant signaling pathway was detected by ELISA. The activation products of the Nrf2 response element were used as the detection index in the experiment; A batch is considered qualified if the ratio of activity level to the positive standard reaches ≥80%. This test serves as a core validation indicator for the "functional expressibility" of brewer's yeast peptides and is suitable for batch-to-batch consistency assessment.

[0067] This quality control method covers both physicochemical properties (molecular weight) and assesses the correlation between functional mechanisms (Nrf2 pathway activation), providing a standardized and objective evaluation method for brewer's yeast peptides in functional food development, quality supervision, and registration applications.

[0068] In summary, the results above indicate that brewer's yeast peptides can effectively address age-induced sarcopenia through a dual mechanism of antioxidation and anti-inflammation. At the functional level, it can improve the muscle endurance and grasping ability of mice; At the tissue level, it promotes type I collagen expression and improves muscle structure; At the molecular level, it enhances the activity of antioxidant enzymes and reduces the level of inflammatory factors.

[0069] In addition, brewer's yeast peptides have good safety, oral adaptability and industrial preparation potential. They can be widely used as core active ingredients in the development of functional food products such as anti-aging nutritional supplements, sarcopenia-specific foods and sports rehabilitation formulas, and have good market prospects and application value.

[0070] It should be understood that the above specific embodiments are only used to illustrate the technical principles and preferred embodiments of the present invention, and are not intended to limit the scope of protection of the present invention. Without departing from the spirit and essence of the present invention, those skilled in the art can make various equivalent substitutions, combinations, improvements or derivative modifications to the technical solutions of the present invention, and these should all be considered to fall within the scope of protection claimed by the present invention.

[0071] The claims of this invention are drafted based on the technical features described in the specification. The functional descriptions, technical parameters, methods of use, and implementation effects involved have been fully disclosed in this specification and possess both disclosure and feasibility. The scope of protection of the claims shall be determined by the content of the claims, and the specification and drawings may be used to interpret the claims.

Claims

1. Use of a beer yeast peptide for the preparation of a composition for improving muscle loss, characterized in that: The beer yeast peptide promotes muscle repair, relieves muscle protein degradation, and thus plays a role in improving muscle reduction by the following dual mechanisms: (a) Antioxidant mechanism: by increasing the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in muscle tissue and serum, reducing malondialdehyde (MDA) levels, and enhancing antioxidant capacity to reduce oxidative damage to muscle fibers; (b) Anti-inflammatory mechanism: by inhibiting the expression of inflammatory factors interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α), reducing chronic inflammation levels to block inflammation-mediated muscle reduction.

2. Use according to claim 1, characterized in that: The beer yeast peptide is prepared by the following steps: (1) Using beer yeast, one of the beer brewing products, as the raw material; (2) Enzymatic hydrolysis with a complex protease, including trypsin and flavor protease, at pH 7.0 and 50°C for 4 hours, and then terminating the reaction by pasteurizing at 95°C for 10 minutes; (3) Purification by membrane separation and other means to obtain peptide fragments with a relative molecular mass of less than 5000 Da.

3. Use according to claim 1 or 2, characterized in that: The muscle reduction is sarcopenia associated with aging, and the composition can be used to improve at least one of the following muscle function indicators: (a) In the fatigue rotarod test, compared with the D-galactose-induced aging model group, the time on the rod is extended by no less than 4 minutes, and the test conditions are 20 rpm and a 5 cm ground clearance; (b) In the four-limb grip strength test, compared with the D-galactose-induced aging model group, the grip strength maintenance time is increased by no less than 1 minute, and the test conditions are a 2 mm diameter metal wire and a 30° inclination angle.

4. Use according to claim 3, characterized in that: The composition improves muscle tissue structure by increasing the content of type I collagen, with the integrated optical density value IOD of type I collagen in the tibialis anterior muscle or gastrocnemius muscle being increased by more than 20% compared with the control group.

5. Use according to any one of claims 1 to 4, characterized in that: The dosage of the beer yeast peptide for old animals is 50-200 mg / kg / day, and the administration is performed by the oral route.

6. A functional food for improving sarcopenia, characterized by, The food comprises the following components: (a) The beer yeast peptide of claim 2; (b) A pharmaceutically or food-acceptable carrier; The dosage form of the functional food is powder, tablet, or liquid beverage.

7. The functional food according to claim 6, characterized in that: The beer yeast peptide accounts for 10%-90% of the total weight in the composition, and its quality indicators include: (a) The total protein content is not less than 80%, and the detection method is the Kjeldahl nitrogen determination method; (b) The antioxidant activity EC50 is not higher than 2 mg / mL, and the detection method is the DPPH method.

8. A test method for evaluating the improvement effect of muscle decrease, characterized by, The following steps are included: (1) Determine the change in lean meat / fat ratio by a body composition analyzer, wherein an increase in lean meat ratio of more than 5% is determined to be effective; (2) Detect the integrated optical density value IOD of type I collagen and type III collagen in muscle tissue using the Sirius red staining method, wherein an increase in IOD value of more than 15% is determined to be effective.

9. A method for quality control of beer yeast peptides, characterized in that, The following detection methods are used to confirm its activity and consistency: (a) Use liquid chromatography-mass spectrometry (LC-MS) to detect the molecular weight distribution, with the main peak concentrated in the range of 300-1000 Da; (b) using enzyme-linked immunosorbent assay (ELISA) to detect the Nrf2 pathway activation ability, and the activation level is not less than 80% under the standard product control.

10. Use, composition or method according to any one of claims 1 to 9, characterized in that: The beer yeast peptide changes the muscle mass, improves the exercise capacity, and realizes the promotion effect on the exercise capacity of the old animals through the antioxidant mechanism and the anti-inflammatory mechanism in claim 1.

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