Green tea extract, hydrolyzed keratin and vitamin C composition for delaying senescence of cats and dogs
By combining green tea extract, hydrolyzed keratin, and vitamin C, the side effects of existing anti-aging drugs have been addressed, achieving safe and effective anti-aging effects in cats and dogs, restoring cell function and delaying aging.
Patent Information
- Application Number
- CN202511315350.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-09-15
- Publication Date
- 2025-12-19
AI Technical Summary
Existing anti-aging drugs have side effects when used on cats and dogs, and their long-term safety remains to be verified. There is a need to develop safer and more effective anti-aging strategies.
A combination of green tea extract, hydrolyzed keratin, and vitamin C is used to prepare tablets, capsules, powders, granules, or oral liquids through a specific process for nutritional intervention and health maintenance in cats and dogs.
It significantly slows down the aging process in cats and dogs, restores cell proliferation capacity, protects cells from oxidative damage, and achieves anti-aging effects by regulating the Hippo pathway.
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Figure CN121154786A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application belongs to the technical field of pet anti-aging food preparation, and specifically relates to the use of green tea extract, hydrolyzed keratin and vitamin C for anti-aging of cat mammary gland cells and dog mammary gland cells. BACKGROUND
[0002] Cellular senescence is a complex process, the nature of which was first revealed by Hayflick and Moorhead in 1961. Researchers found that normal fibroblasts cultured in vitro would inevitably enter a growth-arrested state after a certain number of divisions. This process marked the entry of cells into the senescence phase, during which their physiological functions gradually declined and they lost the potential to continue to proliferate. However, it is worth noting that although these senescent cells have reduced functionality, they still maintain a certain level of activity and can have a profound impact on the function of surrounding tissues for a long time. Currently, a variety of anti-aging drugs have been developed, including eight-son capsules, rapamycin, and beta-nicotinamide mononucleotide. These drugs can delay the aging process to some extent, but they also have certain side effects, such as immune system suppression, gastrointestinal discomfort, etc. caused by rapamycin, and the long-term safety still needs further research. Therefore, in the future, it is still necessary to continuously develop more effective and safe anti-aging drugs to meet the needs of cats and dogs in the aging process to address health problems or geriatric syndromes. SUMMARY
[0003] The purpose of the present application is to provide a new strategy for the treatment of health problems or geriatric syndromes in cats and dogs during the aging process, and it is found that green tea extract, hydrolyzed keratin, and vitamin C have obvious anti-aging effects on cat mammary gland cells and dog mammary gland cells. The method is simple to operate and low in cost.
[0004] The present application provides a green tea extract, hydrolyzed keratin, and vitamin C composition for delaying the aging of cats and dogs, wherein the mass fraction of each component is as follows: green tea extract: 1-3 parts;
[0005] Hydrolyzed keratin: 10-30 parts;
[0006] Vitamin C: 1-2 parts.
[0007] The components in the composition synergistically delay aging and are suitable for nutritional intervention and health maintenance of pet animals such as cats and dogs.
[0008] The green tea extract is prepared according to the following steps: green tea leaves are dried and crushed, the crushed product is added into water at 100-300 g / L, and then extracted at 60-80 DEG C for 1-2 h, the filtrate is reserved after filtration, and the same volume of water is added into the residue, and then extracted again at 60-80 DEG C for 1-2 h, the filtrate is combined after filtration, and then concentrated, and the concentrate is dried and crushed to obtain the green tea extract.
[0009] The hydrolyzed keratin is prepared according to the following steps: wool is washed with water and alcohol to remove dust, dirt and grease and other impurities on the hair, and then dried and crushed, the crushed product is added into sodium sulfite solution (10-50 g / L) at 100-200 g / L, and then soaked for 1-2 h after adjusting pH to 9-11, and then the whole is transferred into a high-temperature and high-pressure device, and then treated at 120-140 DEG C and 0.1-0.15 MPa for 30-60 min, and then distilled water is added into the product after cooling, and then heated to 40-70 DEG C, and then adjusted to pH 9-11, and then keratinase (50000 U / g) is added, and the mass of the keratinase is 0.5-1% of the mass of the crushed product, and then the product is stirred for 1-3 h at constant temperature, and then boiled in water bath for 10-15 min to inactivate the enzyme, and then the filtrate is reserved after centrifugation, and then adjusted to pH 6.0-7.0, and then concentrated, and then spray-dried to obtain the hydrolyzed keratin.
[0010] The application provides applications of green tea extract, hydrolyzed keratin and vitamin C in promoting in-vitro proliferation of cat mammary gland cells and dog mammary gland cells.
[0011] The application provides applications of green tea extract, hydrolyzed keratin and vitamin C in partially restoring in-vitro proliferation ability of cat mammary gland cells and dog mammary gland cells which is reduced due to dactinomycin.
[0012] The application provides applications of green tea extract, hydrolyzed keratin and vitamin C in restoring up-regulation of marker protein levels of cat mammary gland cells and dog mammary gland cells which is induced by dactinomycin.
[0013] The application provides applications of green tea extract, hydrolyzed keratin and vitamin C in protecting cat mammary gland cells and dog mammary gland cells from oxidative damage.
[0014] The application provides applications of green tea extract, hydrolyzed keratin and vitamin C in regulating Hippo pathway to realize anti-aging effect on cat mammary gland cells and dog mammary gland cells.
[0015] The application discloses applications of green tea extract, hydrolyzed keratin and vitamin C in preparing medicines for promoting in-vitro proliferation of cat mammary gland cells and dog mammary gland cells.
[0016] The application discloses applications of green tea extract, hydrolyzed keratin and vitamin C in preparing anti-aging medicines for cat mammary gland cells and dog mammary gland cells.
[0017] The application discloses application of green tea extract, hydrolyzed keratin and vitamin C in preparation of a drug for protecting feline mammary gland and canine mammary gland cells from oxidative damage.
[0018] The application discloses application of a composition of green tea extract, hydrolyzed keratin and vitamin C to related indexes such as exercise capacity and oxidative stress of a fast-aging mouse.
[0019] The dosage form of the above-mentioned pharmaceutical composition can be a tablet, a capsule, a powder, a granule or an oral liquid agent.
[0020] In some embodiments, it is found in an in-vitro experiment that the green tea extract, the hydrolyzed keratin and the vitamin C promote in-vitro proliferation of feline mammary gland and canine mammary gland cells.
[0021] In some embodiments, it is found in an in-vitro experiment that the green tea extract, the hydrolyzed keratin and the vitamin C can partially restore the decreased in-vitro proliferation ability of feline mammary gland and canine mammary gland cells caused by actinomycin D.
[0022] In some embodiments, it is found in an in-vitro experiment that the green tea extract, the hydrolyzed keratin and the vitamin C can restore the protein level up-regulation of a senescence marker in feline mammary gland and canine mammary gland cells induced by actinomycin D.
[0023] In some embodiments, it is found in an in-vitro experiment that the green tea extract, the hydrolyzed keratin and the vitamin C have a significant effect on oxidative damage protection of feline mammary gland and canine mammary gland cells.
[0024] In some embodiments, it is found in an in-vitro experiment that the anti-aging effect of the green tea extract, the hydrolyzed keratin and the vitamin C on feline mammary gland and canine mammary gland cells is regulated by the Hippo pathway.
[0025] In some embodiments, the effect of the composition of the green tea extract, the hydrolyzed keratin and the vitamin C on related indexes such as exercise capacity and oxidative stress of a fast-aging mouse is the effect of delaying aging of the fast-aging mouse.
[0026] Compared with the prior art, the application has the following advantages:
[0027] 1) The green tea extract is an active ingredient extracted from green tea leaves and has multiple effects and functions. It is found in the present research that the green tea extract has an anti-aging effect on feline mammary gland and canine mammary gland cells.
[0028] 2) The hydrolyzed keratin can be eaten by animals after special treatment and provides a high-quality protein source for the animals. It is found in the present research that the hydrolyzed keratin has a new function and provides a new strategy for treatment of a senile syndrome occurring in the aging process of cats and dogs.
[0029] 3) Vitamin C is a clinical drug, and the present study found that it has a new function and is low in price, which belongs to "old drug new use".
[0030] 4) The composition of green tea extract, hydrolyzed keratin, and vitamin C has an effect of delaying the aging of rapidly aging mice. BRIEF DESCRIPTION OF DRAWINGS
[0031] Figure 1 Growth line graph of green tea extract, hydrolyzed keratin, and vitamin C promoting the proliferation of cat mammary gland and dog mammary gland cells in vitro.
[0032] Figure 2 Growth line graph of green tea extract, hydrolyzed keratin, and vitamin C being able to partially restore the decreased proliferation ability of cat mammary gland and dog mammary gland cells in vitro caused by actinomycin D.
[0033] Figure 3 Growth line graph of green tea extract, hydrolyzed keratin, and vitamin C being able to restore the up-regulation of protein levels of aging markers in cat mammary gland and dog mammary gland cells induced by actinomycin D.
[0034] Figure 4 Growth line graph of green tea extract, hydrolyzed keratin, and vitamin C having a significant effect on the oxidative damage protection of cat mammary gland and dog mammary gland cells.
[0035] Figure 5 Growth line graph of green tea extract, hydrolyzed keratin, and vitamin C on the anti-aging effect of cat mammary gland and dog mammary gland cells being regulated by the Hippo pathway. DETAILED DESCRIPTION
[0036] The present application will be further described by way of examples, but the present application is not limited to the scope of the examples. The experimental methods in the following examples are not specified, and the methods are selected according to conventional methods and conditions, or according to the instructions of the product.
[0037] The green tea extract described in the present application is prepared according to the following steps: after drying the green tea leaves, the leaves are crushed, and the crushed material is added to water at 100-300 g / L, and then extracted at 60-80°C for 1-2 hours. After filtration, the filtrate is collected, and an equal volume of water is added to the residue, which is then extracted again at 60-80°C for 1-2 hours. After filtration, the filtrates are combined and concentrated, and the concentrate is dried and crushed to obtain the green tea extract.
[0038] The hydrolyzed keratin according to the present application is prepared by the following steps: taking wool, removing dust, dirt and grease and other impurities on the hair by water washing and alcohol washing, drying and crushing. The crushed material is added into sodium sulfite solution (10-50 g / L) at 100-200 g / L, and soaked for 1-2 h after adjusting pH to 9-11. Then the whole is transferred into a high temperature and high pressure equipment, and treated under the conditions of temperature 120-140 ℃ and pressure 0.1-0.15 MPa for 30-60 min. After cooling, an equal volume of distilled water is added, and the temperature is raised to 40-70 ℃, the pH is adjusted to 9-11, and keratinase (50000 U / g) is added, the mass of keratinase is 0.5-1% of the mass of the crushed material. After constant temperature stirring and enzymatic hydrolysis for 1-3 h, the enzyme is inactivated in a boiling water bath for 10-15 min. After centrifugation, the filtrate is obtained, the pH is adjusted to 6.0-7.0, and the hydrolyzed keratin is prepared by spray drying after concentration.
[0039] Figure 1 shows the growth line graph of green tea extract, hydrolyzed keratin and vitamin C promoting the in vitro proliferation of cat mammary gland and dog mammary gland cells. Figure 1 ) of Example 1.
[0040] Different concentrations of green tea extract, hydrolyzed keratin and vitamin C were added to the culture medium of PETCC#2478 (dog mammary gland) cells and PETCC#81 (cat mammary gland) cells for 24 h, and the OD value of the cell culture medium was detected by CCK8 kit, as shown in Figure 1 .
[0041] The results show that green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C has stronger ability to promote the in vitro proliferation of PETCC#2478 (dog mammary gland) cells and PETCC#81 (cat mammary gland) cells.
[0042] Figure 2 shows the growth line graph of green tea extract, hydrolyzed keratin and vitamin C partially restoring the decrease in the in vitro proliferation ability of cat mammary gland and dog mammary gland cells caused by actinomycin D. Figure 2 ) of Example 2.
[0043] Actinomycin D (5 μg / mL) was added to the culture medium of PETCC#2478 (dog mammary gland) cells and PETCC#81 (cat mammary gland) cells, and green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) was added at the same time for 24 h, and the OD value of the cell culture medium was detected by CCK8 kit, as shown in Figure 2 .
[0044] The results show that green tea extract, hydrolyzed keratin and vitamin C can partially restore the decrease in the in vitro proliferation ability of cat mammary gland and dog mammary gland cells caused by actinomycin D.
[0045] Example 3 Green tea extract, hydrolyzed keratin, vitamin C can restore the protein level up-regulation of senescence markers in cat mammary and dog mammary cells induced by actinomycin D (as shown in Figure 3 ).
[0046] Green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) was added to the culture medium of PETCC#2478 (dog mammary) cells and PETCC#81 (cat mammary) cells for 24 hours to collect protein samples, and the expression levels of p53 and p21 in the cells were detected by Western blot.
[0047] Actinomycin D (5 μg / mL) was added to the culture medium of PETCC#2478 (dog mammary) cells and PETCC#81 (cat mammary) cells, and green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) was added at the same time for 24 hours to collect protein samples, and the expression levels of p53 and p21 in the cells were detected by Western blot.
[0048] The results show that green tea extract, hydrolyzed keratin, vitamin C can restore the protein level up-regulation of senescence markers in cat mammary and dog mammary cells induced by actinomycin D.
[0049] Example 4 Green tea extract, hydrolyzed keratin, vitamin C have significant effects on the protection of oxidative damage in cat mammary and dog mammary cells (as shown in Figure 4 ).
[0050] H2O2 (500 μmol / L) was added to the culture medium of PETCC#2478 (dog mammary) cells and PETCC#81 (cat mammary) cells, and green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) was added at the same time for 24 hours to collect protein samples, and the expression levels of p53 and p21 in the cells were detected by Western blot.
[0051] The results show that green tea extract, hydrolyzed keratin, vitamin C have significant effects on the protection of oxidative damage in cat mammary and dog mammary cells.
[0052] Example 5 The anti-aging effect of green tea extract, hydrolyzed keratin, vitamin C on cat mammary and dog mammary cells is regulated by the Hippo pathway (as shown in Figure 5 ).
[0053] The expression levels of YAP and P-YAP in cells were detected by Western blotting after adding green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) to the culture medium of PETCC#2478 (dog mammary gland) cells and PETCC#81 (cat mammary gland) cells for 24 hours to collect protein samples.
[0054] The expression levels of YAP and P-YAP in cells were detected by Western blotting after adding actinomycin D (5 μg / mL) to the culture medium of PETCC#2478 (dog mammary gland) cells and PETCC#81 (cat mammary gland) cells, and adding green tea extract (200 μg / mL) / hydrolyzed keratin (20 mg / mL) / vitamin C (3 mmol / L) for 24 hours to collect protein samples.
[0055] The results show that the anti-aging effects of green tea extract, hydrolyzed keratin, and vitamin C on cat mammary gland and dog mammary gland cells are regulated by the Hippo pathway.
[0056] Example 6 Effect of combination of green tea extract, hydrolyzed keratin, and vitamin C on delaying the aging of fast-aging mice.
[0057] Test animals and grouping: male SAMP8 fast-aging mice, body weight 28-32 g. The mice were fed in four groups, the control group (normal diet: the mice were fed with standard normal diet every day, each mouse ingested 7 g of commercially available standard full-price mouse feed per day, and the water was free, without adding any functional ingredients), experimental group 1 (supplemented with vitamin C 0.01 g, green tea extract 0.01 g, and hydrolyzed keratin 1 g based on the normal diet), experimental group 2 (supplemented with vitamin C 0.02 g, green tea extract 0.02 g, and hydrolyzed keratin 2 g based on the normal diet), and experimental group 3 (supplemented with vitamin C 0.01 g, green tea extract 0.03 g, and hydrolyzed keratin 3 g based on the normal diet), 5 mice per group. The test was conducted for 28 days. The normal diet was 7 g of commercially available mouse feed per mouse per day, and the water was free.
[0058] Test method:
[0059] 1) Body weight and feed intake recording
[0060] The body weight and feed intake of each group of mice were recorded before and after the feeding test for 28 days, and the change trend of the material before and after feeding was analyzed.
[0061] 2) Respiratory rate test
[0062] Respiratory rate test was conducted before and 28 days after the feeding experiment. The mice were allowed to familiarize with the test environment before the experiment to prevent stress. After the mice's breathing was stable, the oxygen consumption of the mice was determined using a respiratory rate tester.
[0063] 3) Rotarod fatigue test
[0064] Physical fitness test was conducted before and 28 days after the feeding experiment. Before starting the test, each mouse was placed on the rotating rod to train at a slow constant speed. In the formal experiment, the mouse was placed on the rod, facing away from the direction of rotation, and had to walk forward to stay on the rod; the motor was started and the timer was started. When the animal fell off the rod, the test ended. When the mouse fell, the time on the rod indicated by the timer was recorded.
[0065] 4) Oxidative stress (SOD, MDA)
[0066] Before and 28 days after the feeding experiment, the mice's blood was periodically collected, the serum was separated, and the test was conducted according to the kit method.
[0067] The results showed:
[0068] 1) Mouse weight and food intake
[0069] The changes in the body weight of mice in each group before and after feeding the material are shown in Table 1. After 28 days of feeding the sample, the average increase in body weight of mice in the control group was 11.22%, while that in experimental groups 1, 2 and 3 was 13.36%, 17.62% and 18.00%, respectively, which was 1.19, 1.57 and 1.60 times that of the control group.
[0070] Table 1 Body weight (g) of mice before and after feeding the material
[0071]
[0072] # Significant difference compared with the control group, p<0.05
[0073] The changes in the food intake of mice in each group before and after feeding the material are shown in Table 2. After 28 days of feeding the sample, there was no significant difference in food intake between the experimental and control groups, indicating that the gavage of the material did not affect the palatability of the mouse feed and the feeding habits of the mice. Combined with the analysis of other functional indicators of the fast-aging mice, it can be inferred that the differences in the physiological functions of the mice were not caused by differences in feeding.
[0074] Table 2 Food intake (g) of mice before and after feeding the material
[0075]
[0076] # Significant difference compared with the control group, p<0.05
[0077] 2) Mouse respiration rate test
[0078] The results of the mouse respiration rate test before and after feeding the material are shown in Table 3. After 28 days of feeding the material, the respiration rate of the control group mice decreased by 8.31%. The respiration rates of the experimental groups 1, 2, and 3 increased by 3.91%, 8.06%, and 8.73%, respectively. Among them, the experimental groups 2 and 3 had the most significant effect on increasing the respiration rate of mice.
[0079] Table 3: Respiration rate of fast-aging mice before and after feeding the material (ppm / g / min)
[0080]
[0081] # Significant difference compared with the control group, p < 0.05
[0082] 3) Mouse rotarod fatigue test
[0083] The results of the mouse rotarod fatigue test before and after feeding the material are shown in Table 4. After 28 days of feeding the material, the rotarod time of the control group mice decreased by 1.08%. The rotarod times of the experimental groups 1, 2, and 3 increased by 10.93%, 12.65%, and 24.23%, respectively. This indicates that feeding the material of the experimental group 3 can improve the physical fitness of mice and alleviate exercise-induced fatigue.
[0084] Table 4: Rotarod time of fast-aging mice before and after feeding the material (min)
[0085]
[0086] # Significant difference compared with the control group, p < 0.05
[0087] 4) Mouse oxidative stress level
[0088] The contents of superoxide dismutase (T-SOD) and malondialdehyde (MDA) in the serum of fast-aging mice before and after feeding the material are shown in Table 5. After 28 days of feeding the material, the contents of T-SOD in the serum of the experimental groups 1, 2, and 3 increased by 1.15, 1.14, and 1.25 times compared to the control group, respectively. Among them, the content of T-SOD in the serum of the experimental group 3 increased the most significantly. After 28 days of feeding the material, the contents of MDA in the serum of the experimental groups 1, 2, and 3 decreased by 22.79%, 29.54%, and 39.66% compared to the control group, respectively. Among them, the content of MDA in the serum of the experimental group 3 decreased the most significantly. This indicates that feeding the material of the experimental group 3 can effectively alleviate the oxidative stress caused by aging in mice and improve the body's resistance to oxygen free radicals.
[0089] Table 5: Oxidative stress level of fast-aging mice before and after feeding the material
[0090]
[0091] Significant difference compared with the control group, p<0.05.
Claims
1. A composition of green tea extract, hydrolyzed keratin, and vitamin C for delaying aging in cats and dogs, characterized in that... Of which, by mass fraction: green tea extract: 1-3 parts; Hydrolyzed keratin: 10-30 parts; Vitamin C: 1-2 servings.
2. The composition of green tea extract, hydrolyzed keratin, and vitamin C for delaying aging in cats and dogs according to claim 1, characterized in that... The green tea extract is extracted according to the following steps: dried green tea leaves are pulverized, and the pulverized material is added to water at a concentration of 100-300 g / L. The mixture is then extracted at 60-80°C for 1-2 hours. After filtration, the filtrate is collected. An equal volume of water is added to the residue, and the mixture is extracted again at 60-80°C for 1-2 hours. After filtration, the filtrates are combined and concentrated. The concentrate is dried and pulverized to obtain the green tea extract.
3. The composition of green tea extract, hydrolyzed keratin, and vitamin C for delaying aging in cats and dogs according to claim 1, characterized in that... The hydrolyzed keratin is extracted according to the following steps: Wool is washed with water and alcohol to remove dust, dirt, grease, and other impurities, then dried and pulverized. The pulverized material is added at a concentration of 100–200 g / L to a sodium sulfite solution of 10–50 g / L, the pH is adjusted to 9–11, and the mixture is soaked for 1–2 hours. Then, the entire mixture is transferred to a high-temperature, high-pressure (HTHP) device and treated at 120–140°C and 0.1–0.15 MPa for 30–60 minutes. After cooling, an equal volume of distilled water is added, the temperature is raised to 40–70°C, the pH is adjusted to 9–11, and keratinase is added at a concentration of 0.5–1% of the pulverized material. Enzymatic hydrolysis is carried out at a constant temperature with stirring for 1–3 hours, followed by enzyme inactivation in a boiling water bath for 10–15 minutes. After centrifugation, the filtrate is collected, the pH is adjusted to 6.0–7.0, concentrated, and then spray-dried to obtain the hydrolyzed keratin.
4. The use of the green tea extract, hydrolyzed keratin, and vitamin C composition for delaying aging in cats and dogs according to claim 1, characterized in that... A drug used to promote the in vitro proliferation of feline and canine mammary cells.
5. The use of the green tea extract, hydrolyzed keratin, and vitamin C composition for delaying aging in cats and dogs according to claim 1, characterized in that... Drugs used to delay the aging of mammary cells in cats and dogs.
6. The use of the green tea extract, hydrolyzed keratin, and vitamin C composition for delaying aging in cats and dogs according to claim 1, characterized in that... Drugs used to protect against oxidative damage to feline and canine mammary cells.
7. The use of the green tea extract, hydrolyzed keratin, and vitamin C composition for delaying aging in cats and dogs according to claim 1, characterized in that... A nutritional supplement used to delay aging in SAMP8 mice.
8. The composition of green tea extract, hydrolyzed keratin, and vitamin C for delaying aging in cats and dogs according to claim 1, characterized in that... The composition includes green tea extract, hydrolyzed keratin, vitamin C, and a pharmaceutically acceptable carrier.
9. The composition of green tea extract, hydrolyzed keratin, and vitamin C for delaying aging in cats and dogs according to claim 1, characterized in that... Its dosage forms include tablets, capsules, powders, granules, or oral liquids.