A hypoglycemic composition containing a seabuckthorn protein peptide and application thereof

Abstract

The application discloses a hypoglycemic composition containing Hippophae rhamnoides L. protein peptides and application thereof, and belongs to the field of compositions. The hypoglycemic composition containing Hippophae rhamnoides L. protein peptides comprises mulberry leaf extract and Hippophae rhamnoides L. protein peptides, and the mass ratio of the two is (1:5) to (5:1). The composition is prepared by combining the mulberry leaf extract and the Hippophae rhamnoides L. protein peptides in a specific ratio, so that the synergistic effect of the two can be achieved, and good blood glucose lowering effect can be achieved in a short time, and the effect is better than that of similar natural ingredient blood glucose lowering products.

Description

Technical Field

[0001] This invention relates to the field of compositions, and more specifically to a hypoglycemic composition containing sea buckthorn protein peptides and its application. Background Technology

[0002] High blood sugar can occur when the body consumes too much sugar, exercises too little, or is affected by genetic factors or medications. As one of the "three highs" (high blood pressure, high blood sugar, and high cholesterol), long-term high blood sugar can cause lesions in various tissues and organs, leading to acute and chronic complications.

[0003] Common treatments for lowering blood sugar, besides improving dietary habits and regulating lifestyle, mainly involve adjunctive therapy with medications. Currently, in clinical treatment, chemically synthesized drugs such as insulin and its analogues, and alpha-glucosidase inhibitors are frequently used. However, these drugs have many side effects; for example, insulin and its analogues may cause short-term hypoglycemia, accompanied by symptoms such as weakness in the limbs and palpitations. Therefore, people are beginning to use natural ingredients with fewer toxic side effects as alternatives. These natural ingredients also possess the property of "medicine and food as a treatment," which can further reduce their harm to the human body.

[0004] However, some existing natural ingredient drugs and raw materials with hypoglycemic effects have single active ingredients, which have very limited effects on lowering blood sugar. Furthermore, long-term use can easily lead to drug resistance, and they still cannot completely replace traditional synthetic chemical drugs for lowering blood sugar. Summary of the Invention

[0005] Based on the deficiencies of existing technologies, the purpose of this invention is to provide a hypoglycemic composition containing sea buckthorn protein peptides. This composition combines mulberry leaf extract and sea buckthorn protein peptides in a specific ratio, which can achieve a synergistic effect between the two and achieve a good hypoglycemic effect in a short period of time. This effect is superior to existing similar natural hypoglycemic products.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0007] A hypoglycemic composition containing sea buckthorn protein peptides, comprising mulberry leaf extract and sea buckthorn protein peptides in a mass ratio of (1:5) to (5:1).

[0008] Mulberry leaf extract is rich in alkaloids, polysaccharides, and other active substances that have excellent hypoglycemic effects. These active substances can protect pancreatic β-cells and promote insulin synthesis and secretion, thus lowering blood sugar. They can also inhibit the activity of α-glucosidase, thereby suppressing the rise in postprandial blood glucose, improving the postprandial blood glucose curve, and effectively reducing blood sugar. Sea buckthorn itself contains polysaccharides, proanthocyanidins, and phenolic acids, all of which have been proven to have hypoglycemic effects. Furthermore, enzymatically hydrolyzed sea buckthorn protein peptides can also significantly reduce blood glucose levels.

[0009] Through experiments, the inventors discovered that, under specific additive ratios, products prepared from a combination of mulberry leaf extract and sea buckthorn protein peptides exhibit a significant synergistic effect, effectively lowering blood sugar levels in animals, demonstrating a high degree of synergy. Conversely, improper ratios of the two components can lead to antagonistic effects, failing to achieve the synergistic effect.

[0010] Furthermore, compared to other natural ingredients or compositions that also have a hypoglycemic effect, the hypoglycemic composition containing sea buckthorn protein peptides described in this invention has a higher degree of synergy and a more significant therapeutic effect.

[0011] Preferably, the mass ratio of the mulberry leaf extract to the sea buckthorn protein peptide is 1.5:2.5.

[0012] The inventors have verified that when the ratio of the two components in the hypoglycemic composition of this invention is limited to 1.5:2.5, the synergistic effect is the highest, and the effect on reducing blood sugar levels in animals is the best.

[0013] Preferably, the mulberry leaf extract is obtained by sequentially processing mulberry leaves through water extraction, concentration, filtration, and spray drying.

[0014] Preferably, the seabuckthorn protein peptide is prepared from seabuckthorn seed meal.

[0015] Preferably, the sea buckthorn protein peptide can be prepared according to the method of CN114377107A.

[0016] More preferably, the method for preparing the sea buckthorn protein peptide includes the following steps:

[0017] (1) After crushing the sea buckthorn seed meal, disperse it in water. Adjust the pH of the resulting dispersion to 10.0~11.0 and heat to extract. Collect the supernatant, adjust the pH to 5.0~5.5, add phytase for enzymatic hydrolysis, let it stand to precipitate, centrifuge, and obtain precipitate A.

[0018] (2) Dilute precipitate A with water, heat the resulting diluted solution to 80~90℃ and keep it warm, then cool it and adjust the pH to 9.0~9.5, add trypsin for enzymatic hydrolysis, and then subject the hydrolysate to enzyme inactivation, centrifugation, nanofiltration, sterilization and drying to obtain the sea buckthorn protein peptide.

[0019] More preferably, in step (1), the mass ratio of sea buckthorn seed meal to water is 1:(10~11).

[0020] More preferably, in step (1), the temperature of the dispersion extraction is 40~50℃ and the time is 3~4h.

[0021] More preferably, in step (1), the mass ratio of sea buckthorn seed meal to phytase during enzymatic hydrolysis is 1:(0.0004~0.006).

[0022] More preferably, in step (2), the mass ratio of sea buckthorn seed meal to pancreatic enzyme is 1:(0.003~0.005).

[0023] Further verification by the inventors revealed that the active components of the sea buckthorn protein peptides obtained by enzymatic hydrolysis with different amounts of phytase and pancreatic enzyme also differed. In the hypoglycemic composition described in this invention, the sea buckthorn protein peptides prepared by enzymatic hydrolysis with the above-mentioned added amounts of phytase and pancreatic enzyme exhibited the best synergistic effect with mulberry leaf extract.

[0024] Another object of the present invention is to provide the application of the said seabuckthorn protein peptide hypoglycemic composition in the preparation of articles with hypoglycemic function.

[0025] Preferably, the concentration of the hypoglycemic composition used in the product is 250~1000 μg / mL.

[0026] Preferably, the product is a pharmaceutical product.

[0027] More preferably, the drug further includes at least one of excipients, antioxidants, and preservatives.

[0028] More preferably, the dosage form of the drug is any one of liquid dosage form, gas dosage form, solid dosage form, and semi-solid dosage form.

[0029] The hypoglycemic composition containing sea buckthorn protein peptides described in this invention has fewer toxic side effects than chemically synthesized hypoglycemic active substances, and also has better efficacy than other similar natural active substances. Therefore, it is very suitable for preparing some human-friendly and highly effective hypoglycemic products, especially pharmaceuticals. Since this hypoglycemic composition itself belongs to the "medicine and food as a whole" type of product, the pharmaceuticals prepared from it have a wider range of applications and are suitable for the prevention, relief and treatment of various types of diabetes or hyperglycemia.

[0030] The beneficial effect of the present invention is that it provides a hypoglycemic composition containing sea buckthorn protein peptides. The composition combines mulberry leaf extract and sea buckthorn protein peptides in a specific ratio, which can achieve the synergistic effect of the two and achieve good hypoglycemic effect in a short time. This effect is superior to existing similar natural hypoglycemic products. Attached Figure Description

[0031] Figure 1 This is an LC-MS total ion chromatogram of cations and anions in the mulberry leaf extract of the hypoglycemic composition described in Example 1 of the present invention.

[0032] Figure 2 This is a graph showing the combined index results of the efficacy tests of the products in each embodiment and comparative example group in Example 2 of the present invention. Detailed Implementation

[0033] To better illustrate the purpose, technical solution, and advantages of this invention, the invention will be further described below with reference to specific embodiments and comparative examples. The purpose of this description is to provide a detailed understanding of the invention, not to limit its scope. All other embodiments obtained by those skilled in the art without inventive effort are within the protection scope of this invention. Unless otherwise specified, the experimental reagents, raw materials, and instruments designed in the embodiments and comparative examples of this invention are all commonly used reagents, raw materials, and instruments.

[0034] Example 1

[0035] An embodiment of the hypoglycemic composition and its application of the present invention is provided. The hypoglycemic composition of this embodiment includes mulberry leaf extract and sea buckthorn protein peptide, with a mass ratio of 1:1.

[0036] The mulberry leaf extract was purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd., and was obtained by sequentially extracting, concentrating, filtering, and spray drying mulberry leaves. The LC-MS total ion chromatogram of this component for both cations and anions is shown below. Figure 1 As shown.

[0037] The preparation method of the sea buckthorn protein peptide includes the following steps:

[0038] (1) After crushing the sea buckthorn seed meal, pass it through a 20-mesh sieve and disperse it with 10 times its weight of water. Adjust the pH of the resulting dispersion to 11.0 with sodium hydroxide and heat it to 45°C for 3 hours (maintain the pH during extraction). Collect the supernatant, adjust the pH to 5.0 with hydrochloric acid, and enzymatically hydrolyze it with phytase (the mass ratio of sea buckthorn seed meal to phytase is 1:0.0005). Allow it to cool naturally and stand for 2 hours to precipitate. Centrifuge to obtain precipitate A.

[0039] (2) Dilute precipitate A with 3 times its weight of water, heat the resulting diluted solution to 85°C and keep it warm for 15 min, then cool it to 50°C and adjust the pH to 9.0, add pancreatic enzyme from pig pancreas (the mass ratio of sea buckthorn seed meal to pancreatic enzyme is 1:0.004) for enzymatic hydrolysis, adjust the pH to 5.0, and then subject the resulting hydrolysate to enzyme inactivation at 85°C for 30 min, centrifugation, nanofiltration (nanofiltration membrane with a pore size of 150~300 Da), sterilization and spray drying to obtain the sea buckthorn protein peptide.

[0040] Example 2

[0041] An embodiment of the hypoglycemic composition and its application described in this invention differs from Example 1 only in that the mass ratio of mulberry leaf extract and sea buckthorn protein peptide is 2.5:1.5.

[0042] Example 3

[0043] An embodiment of the hypoglycemic composition and its application described in this invention differs from Example 1 only in that the mass ratio of mulberry leaf extract and sea buckthorn protein peptide is 5:1.

[0044] Example 4

[0045] An embodiment of the hypoglycemic composition and its application described in this invention differs from Example 1 only in that the mass ratio of mulberry leaf extract and sea buckthorn protein peptide is 1.5:2.5.

[0046] Example 5

[0047] An embodiment of the hypoglycemic composition and its application described in this invention differs from Example 1 only in that the mass ratio of mulberry leaf extract and sea buckthorn protein peptide is 1:5.

[0048] Comparative Example 1

[0049] A hypoglycemic composition differs from Example 1 only in that it contains mulberry leaf extract and sea buckthorn protein peptide in a mass ratio of 25:1.

[0050] Comparative Example 2

[0051] A hypoglycemic composition differs from Example 1 only in that it contains mulberry leaf extract and sea buckthorn protein peptide in a mass ratio of 1:25.

[0052] Comparative Example 3

[0053] A hypoglycemic composition differs from Example 1 only in that the mulberry leaf extract is replaced with sea buckthorn leaf extract provided by Baoderui Health Industry Co., Ltd.

[0054] Comparative Example 4

[0055] A hypoglycemic composition differs from Example 1 only in that the seabuckthorn protein peptide is replaced with seabuckthorn leaf extract provided by Beijing Baoderui Health Industry Co., Ltd.

[0056] Comparative Example 5

[0057] A hypoglycemic composition differs from Example 1 only in that the mulberry leaf extract is replaced with whey protein supplied by Arla Food Ingredients Ltd.

[0058] Comparative Example 6

[0059] A hypoglycemic composition differs from Example 1 only in that the sea buckthorn protein peptide is replaced with whey protein provided by Arla Food Ingredients Ltd.

[0060] Example 1

[0061] To verify the effects of the two components in the hypoglycemic composition of this invention, a factorial design was performed on the product obtained in Example 1 to verify the synergistic effect of the two components. The experimental method is as follows: A sufficient number of normally developed zebrafish embryos (1 hpf) were pre-selected and randomly assigned to 6-well plates, 30 embryos per well. A 2×2 factorial design was used to establish a normal group, a model control group, a mulberry leaf extract group, a sea buckthorn protein peptide group, and an Example 1 group. The buffer water in the 6-well plates was removed without harming the juvenile fish. Then, 3 mL of high-glucose (2.5% sugar content) solution was quickly added to each well to establish the model for 2 days. Then, 3 mL of test sample containing different concentrations and high-glucose embryo culture medium were quickly added to each well of the test group. After incubation in a biochemical incubator at 28.5±0.5℃ for 2 days, blood glucose levels were measured, and the blood glucose reduction rate was calculated. Blood glucose reduction rate % = (model control group - sample group) / model control group * 100. The test samples used in the mulberry leaf extract group of the test substances were mulberry leaf extract diluted with water to a concentration of 500 μg / mL (the concentration of the entire solution in the 6-well plate, the same below), and the test samples used in the sea buckthorn protein peptide group were sea buckthorn protein peptide diluted with water to a concentration of 500 μg / mL. The sources of the two components were the same as in Example 1; the test sample of Example 1 was the hypoglycemic composition obtained in Example 1 diluted with water to a concentration of 1000 μg / mL.

[0062] The test results are shown in Table 1. It can be seen that the glucose level in the model group was significantly higher than that in the normal control group, indicating that the experimental model was successfully established. Compared with the model group, the glucose levels in each test group were significantly lower (p < 0.01), and the glucose decrease rate was significantly higher (p < 0.01). Furthermore, the glucose levels and glucose decrease rates in the Example 1 group were significantly different from those in the other two individual component test groups (p < 0.05). Factorial design ANOVA was performed on the data, and the results are shown in Table 2. It can be seen that the interaction effect between mulberry leaf extract and sea buckthorn protein peptide in the Example 1 group was significant (P < 0.001), indicating that the two components in the Example 1 group did indeed have a synergistic effect.

[0063] Table 1

[0064]

[0065] Note: ** indicates comparison with the model control group, P<0.01; △ indicates comparison with the mulberry leaf extract and sea buckthorn protein peptide groups, P<0.05.

[0066] Table 2

[0067]

[0068] Furthermore, following the above method, the products of Examples 1-5 and Comparative Examples 1-2 were subjected to the same experiments to investigate the effect of the ratio of the two components in the hypoglycemic composition on the efficacy. Simultaneously, for each example group and comparative example group, in addition to the test solution with a concentration of 1000 μg / mL, test solutions with concentrations of 250 μg / mL and 500 μg / mL were also set as parallel groups for testing. The test data were analyzed using CompuSyn software to determine the combination index (CI) and evaluate the synergistic effect between the two components. The results are shown in Table 3 and... Figure 2(Example 1 is denoted as M25H25, Example 2 as M25H15, Example 3 as M25H5, Example 4 as M15H25, Example 5 as M5H25, Comparative Example 1 as M25H1, and Comparative Example 2 as M1H25). The combination index CI < 1 indicates a synergistic effect between the drugs; combined use enhances the efficacy of each individual drug, with a smaller CI value indicating a stronger synergistic effect. CI = 1 indicates an additive effect between the drugs; the combined use is simply a linear summation of the efficacy of each individual drug. CI > 1 indicates an antagonistic effect between the drugs; combined use may actually reduce the efficacy of each drug. It can be seen that, at different concentrations, the glucose levels of each test group were significantly lower than those of the model group (p < 0.01), and the glucose reduction rate was significantly increased (p < 0.01). However, the glucose reduction in the Example 4 group was relatively higher, indicating that the synergistic effect of mulberry leaf extract and sea buckthorn protein peptide was the best under this composition ratio. In the range of the two group ratios (1:5) to (5:1), the combination index CI of each test group was < 1, indicating that the two active substances had a synergistic effect within this ratio range. However, in the Comparative Examples 1 and 2, CI > 1, indicating that the two active substances had an antagonistic effect under this ratio and did not have a synergistic effect.

[0069] Table 3

[0070]

[0071] Note: ** indicates a comparison with the model control group, P < 0.01

[0072] Example 2

[0073] To verify the optimal combination of the two components in the hypoglycemic composition of the present invention, the products of Comparative Examples 3 to 6 were subjected to the same test as in Example 1. The results showed that the glucose levels and glucose reduction rates of the products of Comparative Examples 3 to 6 could not reach the effect of the group in Example 1. This indicates that the mulberry leaf extract and sea buckthorn protein peptide composition of the present invention can only achieve the ideal synergistic effect when the types and ratios of raw materials are optimized. The lack of components, the substitution of components, or improper ratios will all have a negative impact on the performance of the product.

[0074] Table 4

[0075]

[0076] Note: ** indicates a comparison with the model control group, P < 0.01; △ indicates a comparison with Example 1 group, P < 0.05.

[0077] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A hypoglycemic composition containing sea buckthorn protein peptides, characterized in that, It is composed of mulberry leaf extract and sea buckthorn protein peptide, with a mass ratio of (1:5) to (5:1). The mulberry leaf extract is obtained by sequentially processing mulberry leaves through water extraction, concentration, filtration, and spray drying. The seabuckthorn protein peptide is prepared from seabuckthorn seed meal, and the preparation method of the seabuckthorn protein peptide includes the following steps: (1) After crushing the sea buckthorn seed meal, disperse it in water. Adjust the pH of the resulting dispersion to 10.0~11.0 and heat to extract. Collect the supernatant, adjust the pH to 5.0~5.5, add phytase for enzymatic hydrolysis, let it stand to precipitate, centrifuge, and obtain precipitate A. (2) Dilute precipitate A with water, heat the resulting diluted solution to 80~90℃ and keep it warm, then cool it and adjust the pH to 9.0~9.5, add trypsin for enzymatic hydrolysis, and then subject the hydrolysate to enzyme inactivation, centrifugation, nanofiltration, sterilization and drying to obtain the seabuckthorn protein peptide containing seabuckthorn protein peptide.

2. The hypoglycemic composition containing sea buckthorn protein peptides as described in claim 1, characterized in that, The mass ratio of the mulberry leaf extract to the sea buckthorn protein peptide is 1.5:2.

5.

3. The hypoglycemic composition containing sea buckthorn protein peptides as described in claim 1, characterized in that, In step (1), the mass ratio of sea buckthorn seed meal to phytase during enzymatic hydrolysis is 1:(0.0004~0.006).

4. The hypoglycemic composition containing sea buckthorn protein peptides as described in claim 1, characterized in that, In step (2), the mass ratio of sea buckthorn seed meal to trypsin during enzymatic hydrolysis is 1:(0.003~0.005).

5. The use of the hypoglycemic composition containing sea buckthorn protein peptide as described in any one of claims 1 to 4 in the preparation of an article having hypoglycemic function, wherein the article is a pharmaceutical product.

6. The application as described in claim 5, characterized in that, The hypoglycemic composition containing sea buckthorn protein peptides is used in the product at a concentration of 250~1000 μg / mL.

7. The application as described in claim 5, characterized in that, The dosage form of the drug is any one of liquid dosage form, gas dosage form, solid dosage form, or semi-solid dosage form.

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