Application of lactobacillus delbrueckii KSFY08 in preparation of drugs, food or health care products for improving exercise tolerance

Abstract

The invention provides application of lactobacillus delbrueckii KSFY08 in preparation of drugs, food or health care products for improving exercise tolerance, and relates to the technical field of microorganisms. The lactobacillus delbrueckii KSFY08 disclosed by the invention is preserved in the China General Microbiological Culture Collection Center (CGMCC), and the preservation number is CGMCC No.15715. The research of the invention finds that the lactobacillus delbrueckii KSFY08 can significantly improve the exercise tolerance.

Description

Technical Field

[0001] The invention provides application of Lactobacillus delbrueckii KSFY08 in preparing medicines, foods or health products for improving sports endurance, and belongs to the technical field of microorganisms. Background Art

[0002] For a long time, regular and scientific physical activity has been regarded not only as an effective means to prevent different chronic diseases and related complications, but also as a way to treat diseases. Studies have shown that aerobic exercise can significantly improve physiological adaptability by enhancing cardiovascular capacity and energy metabolites. However, with the accelerated pace of life and work, people are often in a state of continuous or high-intensity work and exercise. Energy consumption makes people prone to fatigue. Continuous fatigue, in turn, prompts people to reduce exercise, which leads to a decline in people's physical fitness and a continuous sub-healthy state of fatigue. In addition, intense or excessive exercise can cause cell and muscle damage, break the balance between oxidative stress and antioxidant systems in the body, lead to the generation of sports fatigue, and have a negative impact on the physical health and career of professional athletes. Therefore, it is of great significance to find intervention methods and effective means to improve physical fitness, enhance sports performance, and improve sports fatigue.

[0003] In recent years, the number of studies on probiotics in the field of sports and the types of nutritional supplements containing probiotics have increased significantly. Current literature shows that probiotics that have a positive effect on improving sports performance mainly include Lactobacillus plantarum, Lactobacillus casei, Lactobacillus acidophilus, Bifidobacterium and Streptococcus thermophilus. At the same time, probiotics have a positive effect on human gastrointestinal function and immune regulation. In addition, animal experiments have confirmed that supplementing with specific probiotics can increase the endurance running time of mice to a certain extent, which provides inspiration for improving human sports endurance performance by supplementing probiotics. The Kashgar area in Xinjiang, my country is an area where ethnic minorities live. The integration of multi-ethnic eating habits makes the local naturally fermented yogurt rich in nutritional value and good health care effects, and the fermented microorganisms in yogurt are also characterized by strong resistance and good diversity, which is of great value for development and utilization. Summary of the invention

[0004] In view of this, the object of the present invention is to provide the application of Lactobacillus delbrueckii KSFY08 in the preparation of medicines, foods or health products that improve sports endurance. At present, the lactic acid bacteria in fermented milk from milk sources in my country mainly include other genera such as Lactobacillus, Bifidobacterium, Lactococcus, Enterococcus, Leuconostoc, Streptococcus and Pediococcus. Lactobacillus delbrueckii belongs to the genus Lactobacillus and is one of the representative species in the genus Lactobacillus. At present, 6 subspecies of Lactobacillus delbrueckii have been found, among which the Bulgarian subspecies is mainly isolated from milk sources and applied to dairy fermentation, and is an edible species specified by the National Health Commission of my country. Studies have shown that new fermented soybeans containing Lactobacillus delbrueckii Bulgarian subspecies exhibit significant anti-fatigue activity. Therefore, this study intends to study the anti-fatigue effect and sports performance improvement effect of Lactobacillus delbrueckii subspecies KSFY08 (Lactobacillus delbrueckii subsp.bulgaricus, LDSB-KSFY08) isolated from naturally fermented yogurt in Kashgar, Xinjiang.

[0005] The Lactobacillus delbrueckii KSFY08 of the present invention is deposited in the General Microbiological Center of China National Committee for Microbiological Culture Collection, with the deposit number being CGMCC No.15715.

[0006] At present, there are four main theories about the mechanism of sports fatigue, namely, the theory of body energy depletion, the theory of accumulation of metabolites in the body, the theory of central protection inhibition, and the theory of free radicals. The theory of body energy depletion believes that when the human body exercises, the large consumption of adenosine triphosphate (ATP) in skeletal muscle or the large loss of glycogen reserves leads to a shortage of energy substances and failure to replenish them in time, which leads to a decrease in muscle working capacity and is an important cause of peripheral fatigue. Therefore, it is of practical significance to detect the content of muscle glycogen and liver glycogen in muscle tissue and serum urea nitrogen.

[0007] In addition, the AMPK-SIRT1 axis is highly related to the body's energy exhaustion. Adenylate activated protein kinase (AMPK) is an evolutionarily conserved energy sensor that regulates cellular metabolism. When the cellular energy state is reduced, AMPK is activated to promote the production of ATP and regulate the use of ATP, thereby restoring energy balance and reducing oxidative stress and other potential adverse cellular events. Similarly, silent information regulator T1 (SIRT1) is activated by changes in energy state to promote gene transcription related to metabolic reactions, and its activity is regulated by nicotinamide phosphoribosyltransferase (NAMPT). Studies have shown that when intracellular glucose is limited, the expression of mouse NAMPT in the mouse myoblast C2C12 series increases, and this increase depends on AMPK. At the same time, glucose restriction also increases SIRT1 activity, thus involving AMPK, NAMPT and SIRT1 in nutrient and energy sensing pathways in skeletal muscle. In recent years, oxidative stress has received increasing attention in the study of sports fatigue protection, and it plays an important role in the pathogenesis of fatigue pathology. Muscle contraction during exercise leads to increased levels of reactive oxygen species (ROS) in skeletal muscle. These highly reactive molecules have many harmful effects, such as reduced force generation and increased muscle atrophy. In response to high levels of ROS, the body will produce antioxidant enzymes such as superoxide dismutase (SOD) to resist the damage caused by oxidative stress. Therefore, increasing the expression of SOD in skeletal muscle after exhaustive exercise can help alleviate oxidative stress.

[0008] This study used a mouse aerobic exercise model to comprehensively evaluate the effects and possible mechanisms of LDSB-KSFY08 in improving mouse exercise endurance, alleviating exercise fatigue, and promoting exercise performance through exhaustive running and swimming tests; blood lactate, muscle glycogen, and liver glycogen concentrations; and AMPK-SIRT1 axis-related gene expression. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1 The effect of LDSB-KSFY08 on the body weight of mice;

[0010] Figure 2 The effect of LDSB-KSFY08 on exhaustive running and exhaustive swimming time in mice;

[0011] Figure 3 The effect of LDSB-KSFY08 on the changes of blood lactate concentration after acute exercise in mice;

[0012] Figure 4 The effect of LDSB-KSFY08 on gastrocnemius muscle fibers in mice;

[0013] Figure 5 The effect of LDSB-KSFY08 on HG and MG concentrations in gastrocnemius muscle of mice;

[0014] Figure 6 This is the effect of LDSB-KSFY08 on the mRNA expression levels of AMPK, NAMPT, SIRT1, PGC1-α, SOD1 and SOD2 in the gastrocnemius muscle of mice. DETAILED DESCRIPTION

[0015] In order to make the purpose, technical scheme and advantages of the present invention clearer, the preferred embodiments of the present invention are further described in detail in conjunction with the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0016] 1 Materials and reagents

[0017] Experimental strain: Lactobacillus delbrueckii subsp.Bulgaricus KSFY08 (LDSB-KSFY08) was isolated from naturally fermented yak yogurt in Kashgar, Xinjiang by the Collaborative Innovation Center for Children's Nutrition and Healthy Development. It is currently deposited in the China General Microbiological Culture Collection Center with the deposit number CGMCC NO.15715, the deposit date: May 12, 2018, and the deposit address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0018] Table 1 Experimental materials and reagents

[0019]

[0020]

[0021] 2. Instruments and Equipment

[0022] Table 2 Instruments and equipment

[0023]

[0024]

[0025] 3 Experimental methods

[0026] 3.1 Animal Experiment Design

[0027] 40 6-week-old (27±1g) SPF male ICR mice were purchased from Hunan Slake Jingda Experimental Animal Co., Ltd. [Animal Qualification License No.: SCXK (Xiang) 2019-0004] and kept in a constant temperature and humidity environment with a temperature of 25±2℃ and a humidity of 50±5%. The mice were fed and drunk freely with light and dark alternation every 12h. All animal experiments were carried out in strict accordance with the laboratory animal use regulations and animal ethics experimental requirements. After 7 days of adaptive feeding, they were randomly divided into 4 groups, namely: quiet control group, exercise control group, LDSB-KSFY08 quiet group and LDSB-KSFY08 exercise group. The whole experiment lasted for 7 weeks, of which the 1st to 6th weeks were the running exercise training period and the 7th week was the behavioral test period. During the experimental period, the quiet control group and the exercise control group were gavaged with 0.2mL of sterile saline at the same time every day, and the LDSB-KSFY08 quiet group and LDSB-KSFY08 exercise group were gavaged with 0.2mL of 1×10 8 CFU / kg of LDSB-KSFY08 bacterial suspension. The exercise control group and the LDSB-KSFY08 exercise group underwent 6 weeks of running exercise training. Before the formal training, they ran at a speed of 12m / min for 25min for 1 day to adapt. In the first week, they started at a speed of 12m / min, and the speed increased by 2m / min every week. From the third week, the slope of the treadmill increased by 5°, and the slope was maintained at 10° from the fourth to the sixth week. The running time was 35min per day. In the seventh week, all mice underwent exhaustive swimming test, exhaustive running test and blood lactate concentration measurement.

[0028] 3.2 Exhaustive swimming test in mice

[0029] Before the exhaustive swimming test, mice were subjected to 10 minutes of weighted swimming adaptation training (water temperature 28±1℃, water depth about 40cm), with a weight of about 5% of the mouse's body weight placed at the base of the mouse's tail. Immediately after the end, the mouse's fur was dried with a towel and blown with hot air until completely dry. A formal exhaustive swimming experiment was conducted on the second day after the weighted swimming adaptation training. The experimental conditions were the same as those of the adaptive training experiment. During the experiment, the mouse's limbs were ensured to continue to move. The exhaustion criterion was that the mouse's nostrils were completely submerged in water for 7 seconds and could not float to the surface. The time taken for the mouse to swim to exhaustion was recorded.

[0030] 3.3 Exhaustive running experiment in mice

[0031] The mice were subjected to exhaustion running test using an 8-channel experimental mouse treadmill. The slope was set to 10°, and the treadmill track speed was gradually increased (10 m / min for 5 min, 16 m / min for 10 min, 21 m / min for 10 min, 26 m / min for 10 min, 31 m / min for 10 min, 36 m / min for 10 min, 41 m / min for 10 min, 46 m / min for 10 min) until the mice were exhausted. The exhaustion criterion for mice was that the number of electric shocks reached 10 times, or they could not continue to run forward for 10 seconds. The time taken for the mice to run to exhaustion was recorded.

[0032] 3.4 Determination of blood lactate concentration in mice

[0033] The lactic acid values ​​of mice were measured at rest before exhaustive running, 5 minutes after exhaustion, and 30 minutes after exhaustion. The blood was collected from the tip of the mouse's tail using a blood collection needle that came with the lactate detector. The first drop of blood was wiped off, and the whole blood was drawn with a test strip to read the data on the detector. Before and after each blood test, the blood collection site on the mouse's tail was disinfected with a cotton ball dipped in iodine.

[0034] 3.5 Mouse dissection and sampling

[0035] After all behavioral tests were completed, the mice were fasted but not watered for 12 hours, and then all mice were euthanized and biological samples were collected. The whole blood of the mice was collected and placed at room temperature for 30 minutes before centrifugation at 4000r / min for 15 minutes at 4°C. The upper serum was separated and aspirated and stored at -80°C for later use.

[0036] 3.6 H&E staining analysis of mouse gastrocnemius muscle

[0037] The gastrocnemius muscle of mice was fixed in paraformaldehyde tissue fixative. After the tissue samples were fixed and dehydrated, they were embedded in paraffin. The paraffin-embedded blocks were cut into 4 μm thin slices by a microtome, and then routinely dewaxed and hydrated with gradient ethanol. Subsequently, they were stained with hematoxylin for 2 minutes, differentiated with 1% hydrochloric acid ethanol, and blued with dilute ammonia water. Finally, they were stained with eosin for 15 seconds, dehydrated with gradient ethanol, and sealed after dehydration. The histological changes of the gastrocnemius muscle of mice were observed and analyzed under an electron microscope.

[0038] 3.7 Determination of HG and MG concentrations in mouse gastrocnemius muscle

[0039] According to the instructions of the enzyme linked immunosorbent assay (ELISA) kit, the concentrations of hepatic glycogen (HG) and muscle glycogen (MG) in the gastrocnemius muscle of mice were measured.

[0040] 3.8 Determination of mRNA expression levels of AMPK-STRI1 pathway-related factors in mouse gastrocnemius muscle

[0041] Weigh an appropriate amount of mouse gastrocnemius muscle tissue, add TRIzol for homogenization to extract total RNA, and then use a spectrophotometer to measure the concentration and purity of the extracted total RNA, and reverse transcribe the total RNA into cDNA according to the instructions of the reverse transcription kit. After reverse transcription, prepare the cDNA reaction system in an eight-tube strip, namely 10 μL of Master mix, 1 μL of upstream and downstream primers, 1 μL of template cDNA, and 7 μL of sterile enzyme-free water. The prepared reaction system was amplified and detected using real-time fluorescence quantitative PCR (Quantitative Real-time PCR, qPCR) technology. Amplification conditions: pre-denaturation at 95°C for 5 min, then denaturation at 95°C for 10 s, annealing at 60°C for 30 s, and the whole process was repeated 40 times. After the determination, GAPDH was used as the internal reference gene and 2 -ΔΔCT The relative expression of the target gene was determined by the formula. The primer sequences used in this experiment are shown in Table 3.

[0042] Table 3 Primer sequences

[0043]

[0044] 3.9 Data processing and statistical analysis

[0045] All data were expressed as mean ± standard deviation (SD) and analyzed using IBM SPSS 23.0 statistical software. One-way analysis of variance (ANOVA) and Duncan's multiple range test were used for comparison between groups. The same lowercase letters in ae indicate no significant difference between the two groups, and different lowercase letters indicate significant difference between the two groups (P < 0.05).

[0046] 4 Results and analysis

[0047] 4.1 Effect of LDSB-KSFY08 on the body weight of mice

[0048] To evaluate the effects of long-term exercise training and LDSB-KSFY08 on the growth status of mice, the body weight of mice was measured and recorded once a week. Figure 1As shown in the figure, after the first week of adaptation, there was no significant difference in the weight of mice in each group. Throughout the experimental period, the weight of mice in each group showed a steady upward trend without abnormal fluctuations. At the end of the 7th week of the experiment, the weight of mice in the exercise control group, LDSB-KSFY08 quiet group, and LDSB-KSFY08 exercise group was significantly different from that in the quiet control group (P<0.05). Among them, the weight of mice in the quiet control group was the heaviest (41.13±1.04g), and the weight of mice in the LDSB-KSFY08 exercise group was the lightest (35.24±1.60g). There was no significant difference between the exercise control group and the LDSB-KSFY08 quiet group (P>0.05), but there was a significant difference between the LDSB-KSFY08 exercise group (P<0.05).

[0049] 4.2 Effect of LDSB-KSFY08 on exhaustive exercise time in mice

[0050] Exhaustive exercise is a common indicator for evaluating the exercise endurance of mice, including exhaustive running and exhaustive swimming. Figure 2 As shown in A, there were significant differences in the time taken by mice in each experimental group to reach exhaustion in the exhaustion running test (P<0.05). The overall trend was that the mice in the quiet control group took the shortest time to reach exhaustion, followed by the exercise control group, the LDSB-KSFY08 quiet group, and the LDSB-KSFY08 exercise group. It is worth mentioning that not only was the time taken by the LDSB-KSFY08 exercise group to reach exhaustion significantly higher than that of the exercise control group, but the time taken by the LDSB-KSFY08 quiet group to reach exhaustion was also significantly longer than that of the exercise control group (P<0.05). At the same time, the exhaustion swimming experiment was also used to detect changes in the exercise endurance of mice in each group, and the experimental results were consistent with those of the exhaustion running experiment ( Figure 2 B), the mice in the LDSB-KSFY08 exercise group took the longest time to reach exhaustion, followed by the LDSB-KSFY08 quiet group, the exercise control group, and the quiet control group, and the exhaustion time between the groups was significantly different (P<0.05). The experimental results show that LDSB-KSFY08 can significantly improve the exercise endurance of mice without exercise training, and can be combined with daily aerobic exercise training to effectively improve the exercise endurance of mice.

[0051] 4.3 Effect of LDSB-KSFY08 on changes in blood lactate concentration after excessive exercise in mice

[0052] Lactic acid is an important indicator for evaluating fatigue and aerobic metabolism after exercise. Figure 3As shown in the figure, at the resting moment before exhaustive exercise, there was no significant difference in the blood lactate concentration among the mice in each group (P>0.05), and the blood lactate concentration was generally at a low level. However, 5 minutes after exhaustive exercise, the blood lactate content of mice in each group increased, and the differences between the groups were significant (P<0.05). Among them, the blood lactate concentration of mice in the quiet control group was the highest among all groups, followed by the exercise control group, LDSB-KSFY08 quiet group and LDSB-KSFY08 exercise group. After 30 minutes of exhaustive exercise, the blood lactate concentration of mice in each group decreased, and the differences between the groups were significant (P<0.05). Among them, the blood lactate content of the LDSB-KSFY08 exercise group was the lowest, followed by the LDSB-KSFY08 quiet group and the exercise control group, and the blood lactate content of the quiet control group was the highest. The results showed that regular daily exercise training can help speed up the conversion of blood lactate after excessive exercise and reduce the accumulation of blood lactate. The intake of LDSB-KSFY08 can not only effectively alleviate the increase in blood lactate concentration after excessive exercise in mice, but also accelerate the conversion of blood lactate, reduce blood lactate concentration, and improve exercise fatigue.

[0053] 4.4 Effects of LDSB-KSFY08 on the histology of mouse gastrocnemius muscle

[0054] Depend on Figure 4 It can be seen that no pathological changes such as injury, edema, inflammation, hemorrhage, and necrosis were found in the gastrocnemius tissue of mice in each group, indicating that the 6-week aerobic exercise training volume was moderate and did not cause muscle tissue damage due to excessive exercise, and it can be used as an effective experimental subject for studying the exercise performance of mice. Among them, the gastrocnemius muscle fibers in the quiet control group were mostly type IIB muscle fibers (white muscle fibers, thick fiber cross-section, light staining), and only a very small number of type I muscle fibers (red muscle fibers, thin fiber cross-section, darker staining) and type IIA muscle fibers (intermediate muscle fibers, thick fiber cross-section, darker staining) were seen. Compared with the quiet control group, the content of type I and type IIA muscle fibers in the exercise control group increased, and the number of these two types of muscle fibers was significantly increased in the LDSB-KSFY08 quiet group, while the gastrocnemius in the LDSB-KSFY08 exercise group contained more type I and type IIA muscle fibers. The experimental results show that long-term exercise training helps to increase the number of type I and type IIA muscle fibers related to endurance. Daily intake of LDSB-KSFY08 can not only increase the number of endurance-related muscle fibers in the gastrocnemius muscle of non-exercise individuals, but also significantly increase the exercise tolerance of the gastrocnemius muscle in combination with exercise training.

[0055] Effects of 4.5LDSB-KSFY08 on HG and MG concentrations in gastrocnemius muscle of mice

[0056] As the main energy source for muscle metabolism, high levels of liver glycogen and muscle glycogen reserves can effectively reduce muscle fatigue symptoms. Figure 5As shown in A and B, the HG and MG content in the gastrocnemius of mice in the quiet control group was the lowest, followed by the exercise control group. Compared with the quiet control group, the HG and MG content in the gastrocnemius of the LDSB-KSFY08 quiet group and the LDSB-KSFY08 exercise group was significantly increased (P<0.05). The experimental results show that a six-week aerobic exercise training can significantly increase the HG and MG content in the gastrocnemius after exhaustive exercise. The intake of LDSB-KSFY08 alone can also effectively promote the recovery of HG and MG concentration in the gastrocnemius. LDSB-KSFY08 combined with aerobic exercise training can exert a better effect.

[0057] Effect of LDSB-KSFY08 on mRNA expression levels of AMPK, NAMPT, SIRT1, PGC1-α, SOD1 and SOD2 in gastrocnemius muscle of mice

[0058] To further analyze the effect of LDSB-KSFY08 on improving sports fatigue and enhancing sports performance, the mRNA expression levels of AMPK, NAMPT, SIRT1, PGC1-α, SOD1 and SOD2 in gastrocnemius muscle were detected. Figure 6 As shown. Compared with the quiet control group without exercise training, the mRNA expression levels of AMPK, NAMPT, SIRT1, PGC1-α, SOD1 and SOD2 in the exercise control group and LDSB-KSFY08 and exercise group mice that underwent 6 weeks of aerobic exercise training were significantly increased (P<0.05). At the same time, the LDSB-KSFY08 quiet group mice also showed significantly higher AMPK, NAMPT, SIRT1, PGC1-α, SOD1 and SOD2 mRNA expression levels than the quiet control group. The results show that daily exercise training can improve physical fitness, improve the decrease in energy metabolism caused by acute exercise, and alleviate the symptoms of oxidative stress after excessive exercise. The daily intake of LDSB-KSFY08 can also achieve the effect of daily exercise training. Daily exercise training combined with LDSB-KSFY08 intake can effectively increase exercise endurance and improve exercise performance.

[0059] 5 Conclusion

[0060] Exercise can promote the body's health, improve physical fitness, and maintain a stable body weight. The results of this experiment showed that the weight gain trend of the mice in the exercise control group and LDSB-KSFY08 exercise group was significantly lower than that in the quiet control group ( Figure 1), indicating that daily moderate exercise can effectively improve the rapid weight gain caused by long-term sitting, and the single LDSB-KSFY08 intervention also has the effect of maintaining stable weight gain. In order to evaluate the effect of LDSB-KSFY08 on the exercise endurance of mice, exhaustive running and exhaustive swimming experiments were conducted. The results showed that the intake of LDSB-KSFY08 can significantly improve the exercise endurance of mice, which is manifested in a significant increase in the exhaustion time of running and swimming ( Figure 2 ), and its effect is even better than the effect of daily exercise training on improving the exercise endurance of mice.

[0061] Lactic acid accumulation is one of the causes of muscle fatigue, and lactic acid has always been regarded as a key factor in inducing skeletal muscle fatigue. First, lactic acid accumulation reduces the activity of phosphofructokinase, a key enzyme in glycolysis, and then affects the resynthesis rate of ATP by reducing the rate of glycolysis; secondly, for muscle cells, the accumulation of lactic acid will form an acidic internal environment, reduce the efficiency of muscle work, and affect the physiological process of skeletal muscle contraction and relaxation; finally, excessive lactic acid accumulation will trigger a series of pathological reactions in muscles, which may cause delayed muscle soreness or non-infectious inflammation. The results of this experiment show that the intake of LDSB-KSFY08 can effectively reduce the accumulation of lactic acid in the blood, and help to quickly clear blood lactate after excessive exercise, thereby playing a role in relieving sports fatigue and promoting the improvement of sports endurance. At the same time, Lactobacillus plantarum CCFM1280 and 73L1, which are also probiotics, have also been reported to relieve fatigue by relieving muscle and blood lactic acid accumulation.

[0062] Skeletal muscle is one of the largest organs in the human body, accounting for about 40% to 50% of the body mass. Skeletal muscle plays an important role in maintaining the body's energy balance and daily activities. Skeletal muscle has a high regenerative capacity. The molecular signaling pathways in skeletal muscle can promote the activation, proliferation and differentiation of myoblasts. Moderate physical activity helps to increase the function of skeletal muscle mitochondria, promote the activity of enzymes involved in metabolic pathways in skeletal muscle, and store glycogen in muscle. Skeletal muscle fibers are mainly divided into three categories, from fast to slow contraction speed, namely type I muscle fibers or slow-twitch fibers, type IIA muscle fibers also known as intermediate muscle fibers, and type IIB muscle fibers also known as fast-twitch fibers. Type I muscle fibers are characterized by slow contraction, not easy to fatigue, and high aerobic respiration capacity; type IIA muscle fibers are characterized by fast contraction speed and resistance to fatigue; and type IIB muscle fibers have the characteristics of fast contraction speed and easy fatigue. Studies have found that skeletal muscle fiber types can be induced and transformed into each other to achieve the purpose of improving athletes' athletic performance and even treating muscle-related diseases. The results of this study showed that regular daily exercise training can increase the number of type I and type IIA muscle fibers in the gastrocnemius muscle, thereby promoting exercise endurance. At the same time, the intake of LDSB-KSFY08 can also promote the transformation of muscle fibers in the gastrocnemius muscle of mice into more fatigue-resistant type I and type IIA muscle fibers. Among them, LDSB-KSFY08 combined with daily exercise training can significantly improve the exercise endurance and fatigue resistance of mice through the transformation of muscle fiber types.

[0063] Glycogen is an important energy source to meet the energy needs and stability of physical activities. During continuous or high-intensity exercise, in order to maintain blood sugar balance, muscle glycogen and liver glycogen in the body will be broken down for energy supply. Therefore, the amount of glycogen reserves in the body is positively correlated with the body's anti-fatigue degree. The results of this experiment showed that LDSB-KSFY08 can significantly alleviate the decrease in the concentration of HG and MG in the gastrocnemius muscle after excessive exercise ( Figure 5 A and B), indicating that LDSB-KSFY08 can alleviate exercise fatigue by reducing the degradation of HG and MG in gastrocnemius muscle.

[0064] AMPK and SIRT1 are fuel-sensitive molecules that coexist in cells throughout the evolutionary process. AMPK is involved in the energy metabolism of various substances in the body. When cells are under stress (such as ischemia, hypoxia, exercise, etc.), the AMPK kinase system will be activated, increasing metabolic processes such as glucose transport and fatty acid oxidation, while blocking metabolic processes such as gluconeogenesis, protein synthesis, and lipids. In mammals, NAMPT is responsible for regulating the essential cofactor NAD of SIRT1, and SIRT1 can activate peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), thereby promoting mitochondrial biogenesis. In addition, PGC-1α plays an important regulatory role in fatty acid oxidation, glucose metabolism, and mitochondrial biogenesis. Therefore, regulating the AMPK-SIRT1 axis can help promote mitochondrial biogenesis and energy metabolism during exercise, thereby delaying the occurrence of fatigue and improving exercise endurance. The results of this experiment showed that LDSB-KSFY08 could significantly increase the mRNA expression levels of AMPK, NAMPT, SIRT1 and PGC-1α in the gastrocnemius muscle of mice, indicating that LDSB-KSFY08 could promote the improvement of athletic performance by activating the AMPK-SIRT1 axis.

[0065] In addition, excessive exercise will lead to peroxidation in the body, which in turn causes oxidative stress damage, and preclinical studies have shown that endurance training can increase the activity of key antioxidant enzymes in skeletal muscle, such as SOD1 and SOD2. Consistent with this, in this study, the exercise control group expressed higher levels of SOD1 and SOD2 than the quiet control group, proving that daily aerobic exercise training helps to increase the content of antioxidant enzymes. It is worth noting that mice that only ingested LDSB-KSFY08 without exercise training showed higher levels of SOD1 and SOD2 than the exercise control group, indicating that LDSB-KSFY08 has the effect of increasing antioxidant enzymes in mice.

[0066] In this study, it can be seen that LDSB-KSFY08 can maintain the stable weight growth of mice, prolong the time of exhaustive running and exhaustive swimming, reduce the blood lactate concentration after excessive exercise, increase the content of muscle glycogen and liver glycogen in gastrocnemius muscle, reduce serum creatine kinase and blood urea nitrogen concentration, and activate the mRNA expression levels of mitochondrial genes (AMPK, NAMPT, SIRT1 and PGC1-α) and skeletal muscle antioxidant genes (SOD1 and SOD2). It shows that Lactobacillus delbrueckii subspecies bulgaricus LDSB-KSFY08 plays a role in improving exercise endurance, reducing exercise fatigue, and promoting exercise performance based on the AMPK-SIRT1 axis. It provides a candidate strain for the development of sports-related nutritional supplements. LDSB-KSFY08 combined with daily exercise training can significantly improve the exercise endurance and anti-fatigue ability of mice through the transformation of muscle fiber types. LDSB-KSFY08 can relieve sports fatigue by reducing the decomposition of HG and MG in the gastrocnemius muscle; LDSB-KSFY08 can significantly increase the mRNA expression levels of AMPK, NAMPT, SIRT1 and PGC-1α in the gastrocnemius muscle of mice, indicating that LDSB-KSFY08 can promote the improvement of sports performance by activating the AMPK-SIRT1 axis; LDSB-KSFY08 has the effect of increasing the antioxidant enzymes in mice.

[0067] The embodiments described above are part of the embodiments of the present invention, rather than all of the embodiments. The detailed description of the embodiments of the present invention is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

Claims

1. Application of Lactobacillus delbrueckii KSFY08 in the preparation of medicines, foods or health products for improving sports endurance, characterized in that: The Lactobacillus delbrueckii KSFY08 is deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration, with the deposit number being CGMCC No.15715.

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