Anti-non-enzymatic glycosylation composition and its application, non-enzymatic glycosylation inhibitor

Abstract

The present invention provides an anti-non-enzymatic glycosylation composition and its application, and a non-enzymatic glycosylation inhibitor. The anti-non-enzymatic glycosylation composition is composed of the following raw materials in parts by weight: 30 to 90 parts of sialic acid, 10 to 40 parts of ginkgo leaf extract, 10 to 40 parts of acerola extract, and 0.1 to 10 parts of rosemary extract. The composition of the present invention can achieve an anti-non-enzymatic glycosylation level comparable to that of the prior art by compounding sialic acid with ginkgo leaf extract, acerola extract, and rosemary extract in a specific ratio, while the stability is also improved. Applying it to health foods or cosmetics with anti-aging effects saves costs and is conducive to production and promotion.

Description

Technical Field

[0001] The invention relates to anti-aging products, in particular to an anti-non-enzymatic glycosylation composition and application thereof, and a non-enzymatic glycosylation inhibitor. Background Art

[0002] The non-enzymatic glycosylation (NEG) reaction in the body refers to the process in which the aldehyde or ketone groups of reducing sugars and the free amino groups in macromolecules such as proteins form brown, fluorescent substances called advanced glycosylation end-products (AGEs) by cross-linking or polymerization under the condition of no enzyme catalysis. AGEs are brown or black macromolecules that appear in the dermis and epidermis. As people age, AGEs are deposited in human skin, causing yellowing of the skin and the production of brown spots and age spots. At the same time, AGEs accelerate the aging of the human body and lead to the occurrence of many chronic degenerative diseases, such as diabetes, atherosclerosis, chronic kidney disease, liver disease, neurodegenerative diseases and other diseases.

[0003] Since non-enzymatic glycosylation reactions are widely and slowly carried out in the body, they can lead to reduced protein function and aging, and then cause aging and pathological changes in body tissues. Based on this, the prior art proposes anti-non-enzymatic glycosylation compositions or AGEs inhibition compositions, which inhibit non-enzymatic glycosylation reactions and / or degrade AGEs, thereby slowing down the aging and pathological changes in body tissues.

[0004] For example, the prior art discloses an AGEs inhibition composition, which is composed of the following raw materials: bilberry extract, ginkgo leaf extract, green tea extract, rosemary extract and vitamins. Although the composition can inhibit the generation and degradation of AGEs, the effective active ingredient of the bilberry extract is anthocyanin, which is extremely unstable and is easily affected by various physical and chemical factors (such as temperature, light, metal ions, oxygen, etc.) to cause oxidation, polymerization, degradation, discoloration and other unstable phenomena. Even if stored in a dark room, it is still easy to fade, which is bound to seriously affect the stability and shelf life of the product, and also restrict the dosage form, production process and storage conditions of the product, increase production costs, and is not conducive to the promotion of the final product. Summary of the invention

[0005] In view of the problems existing in the prior art, the present invention provides an anti-non-enzymatic glycosylation composition and its application, and a non-enzymatic glycosylation inhibitor, so as to increase the stability of the composition and its related products without reducing the anti-non-enzymatic glycosylation effect.

[0006] The present invention adopts the following technical solutions:

[0007] The invention provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 30-90 parts of sialic acid, 10-40 parts of ginkgo leaf extract, 10-40 parts of acerola cherry extract and 0.1-10 parts of rosemary extract.

[0008] In order to achieve the purpose of the present invention, the inventors tried to add stabilizers to the formula disclosed in the prior art to increase the stability of the bilberry extract, but the effect was not satisfactory. The inventors also tried to remove the unstable bilberry extract and use other components with better anti-non-enzymatic glycation effects (such as vitamin B 2 ) to replace bilberry extract, but due to the limited intake of vitamin B group, the anti-non-enzymatic glycosylation effect of the obtained composition is still reduced. After a lot of research, the inventors finally unexpectedly found that sialic acid, which is safe to ingest and has anti-non-enzymatic glycosylation effect but not higher than bilberry extract, can be compounded with ginkgo leaf extract, acerola cherry extract and rosemary extract in a specific ratio to achieve a level of anti-non-enzymatic glycosylation effect equivalent to that of the original formula, while the stability is also improved, and there is no need to add various vitamins, which saves costs and is conducive to production and promotion.

[0009] Furthermore, the invention is composed of the following raw materials in parts by weight: 40-70 parts of sialic acid, 10-20 parts of ginkgo leaf extract, 20-30 parts of acerola cherry extract and 0.1-10 parts of rosemary extract.

[0010] In a preferred embodiment of the present invention, the anti-non-enzymatic glycosylation composition is composed of the following raw materials in parts by weight: 55 parts of sialic acid, 13 parts of ginkgo leaf extract, 30 parts of acerola cherry extract and 2 parts of rosemary extract.

[0011] Furthermore, the anti-non-enzymatic glycosylation composition also includes 5 to 10 parts of natto extract.

[0012] The present invention has found that adding a certain amount of natto extract to the above composition can further enhance the anti-non-enzymatic glycosylation effect, delay body aging, and improve the body's detoxification function.

[0013] The present invention also provides the use of the anti-non-enzymatic glycosylation composition in the preparation of health food or cosmetics with anti-non-enzymatic glycosylation related functions.

[0014] Among them, the anti-non-enzymatic glycosylation composition can be used alone as an active ingredient in health food or cosmetics, or can be used in combination with other components with the same efficacy. Health food is a food that is not intended to treat diseases, but can regulate or alleviate the aging and pathological changes of body tissues (including diabetes, atherosclerosis, etc.), and does not cause any acute, subacute or chronic harm to the human body. Cosmetics refer to a broad sense, and can include skin care products, beauty products, etc., and can be in the form of essences, lotions or facial masks.

[0015] The present invention also provides a non-enzymatic glycosylation inhibitor, comprising the above-mentioned anti-non-enzymatic glycosylation composition and an auxiliary material acceptable in health food science.

[0016] Furthermore, the dosage form of the non-enzymatic glycosylation inhibitor is an oral preparation, including common oral dosage forms such as tablets, powders, and oral liquids.

[0017] Furthermore, the daily intake of sialic acid in the non-enzymatic glycosylation inhibitor is not less than 50 mg; according to the regulations on sialic acid, the recommended daily intake of sialic acid is not more than 500 mg.

[0018] The present invention provides an anti-non-enzymatic glycosylation composition, which can achieve an anti-non-enzymatic glycosylation level equivalent to that of the prior art by compounding sialic acid with ginkgo leaf extract, acerola cherry extract and rosemary extract in a specific ratio, and at the same time, the stability is also improved. The composition is applied to health foods or cosmetics with anti-aging effects, which saves costs and is conducive to production and promotion. DETAILED DESCRIPTION

[0019] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of 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.

[0020] The raw materials and auxiliary materials involved in the following examples, unless otherwise stated, can all be purchased commercially. Among them, each extract can be purchased commercially or made in-house. The sources of the raw materials are as follows:

[0021] Ginkgo leaf extract: sold by Xi'an Prius Bioengineering Co., Ltd., with a total flavonoid glycoside content of 24-26%.

[0022] Acerola cherry extract: sold by Xi'an Ruierli Bioengineering Co., Ltd., with a VC content greater than 17%.

[0023] Rosemary extract: sold by naturex.

[0024] Natto extract: obtained by fermenting soybeans with Bacillus subtilis in a shallow tray, and the eluate is spray-dried. The nattokinase content is greater than 2000 Fu / g.

[0025] Sialic acid: N-acetylneuraminic acid monomer obtained by fermentation. For the specific preparation method, refer to CN109627269B.

[0026] Bilberry extract: Originated from Xi'an, contains 20% anthocyanins.

[0027] Example 1

[0028] This embodiment provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 40 parts of sialic acid, 20 parts of ginkgo leaf extract, 20 parts of acerola cherry extract and 10 parts of rosemary extract.

[0029] Example 2

[0030] This embodiment provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 55 parts of sialic acid, 13 parts of ginkgo leaf extract, 30 parts of acerola cherry extract and 2 parts of rosemary extract.

[0031] Example 3

[0032] This embodiment provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 65 parts of sialic acid, 10 parts of ginkgo leaf extract, 20 parts of acerola cherry extract and 5 parts of rosemary extract.

[0033] Example 4

[0034] This embodiment provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 78 parts of sialic acid, 10 parts of ginkgo leaf extract, 10 parts of acerola cherry extract and 2 parts of rosemary extract.

[0035] Example 5

[0036] This embodiment provides an anti-non-enzymatic glycosylation composition, which is composed of the following raw materials in parts by weight: 50 parts of sialic acid, 13 parts of ginkgo leaf extract, 26 parts of acerola cherry extract, 4 parts of rosemary extract and 7 parts of natto extract.

[0037] Comparative Example 1

[0038] This comparative example sets the composition components according to the daily intake of vitamin B and ginkgo leaf extract. This comparative example provides an anti-non-enzymatic glycation composition, which is composed of the following raw materials in parts by weight: vitamin B 1 1.5 portions, vitamin B 2 1.5 servings and vitamin B 61.5 parts, 20 parts of ginkgo leaf extract, 2 parts of rosemary extract, 40 parts of acerola cherry extract, and the rest are supplemented with microcrystalline cellulose to 100 parts.

[0039] Comparative Example 2

[0040] This comparative example provides an anti-non-enzymatic glycation composition, which is composed of the following raw materials, by weight: 49 parts of bilberry extract, 20.5 parts of ginkgo leaf extract, 24 parts of acerola cherry extract, 2 parts of rosemary extract, and vitamin B 1 1.5 portions, vitamin B 2 1.5 servings and vitamin B 6 1.5 servings.

[0041] Test Example 1 Non-enzymatic glycosylation inhibition ability of each component

[0042] 1. Test principle

[0043] This experiment simulates the process of non-enzymatic glycosylation in vivo in vitro to test the ability of the test substance to inhibit the generation of AGEs and evaluate the potential anti-aging and anti-yellowing effects of the sample.

[0044] 2. Test methods

[0045] 2.1 Reagent preparation

[0046] (1) 0.2 mol / L phosphate buffer (PBS, pH = 7.3-7.4): weigh 7.16 g Na 2 HPO 4 12H 2 O was dissolved in deionized water, and a certain amount of penicillin and streptomycin (antibacterial) was added, and then the volume was adjusted to 100 mL with deionized water to obtain 0.2 mol / L Na 2 HPO 4 Solution; weigh 3.12g NaH 2 PO 4 ·2H 2 O was dissolved in deionized water and the volume was adjusted to 100 mL to obtain 0.2 mol / L NaH 2 PO 4 Solution; take 81mL of 0.2mol / L Na 2 HPO 4 solution and 19 mL of 0.2 mol / L NaH 2 PO 4 The solution is mixed.

[0047] (2) Bovine serum albumin solution (BSA solution): Weigh a certain amount of BSA, add PBS to make up to 25 mL, and filter through a 0.45 μm water filter membrane;

[0048] (3) Fructose solution: Weigh a certain amount of fructose and dilute to 25 mL with PBS;

[0049] (4) BSA-fructose reaction solution: Mix 20 mL of BSA solution and 20 mL of fructose solution.

[0050] 2.2 Sample processing

[0051] The samples were diluted to 10% with deionized water and then diluted to the corresponding concentrations for detection.

[0052] 2.3 Control group and preparation

[0053] (1) Negative control

[0054] PBS was selected to replace the test substance and mixed with the reaction solution, which was the negative control group. The negative control group reflects the situation that there is no inhibitory effect on the non-enzymatic glycosylation reaction in the system.

[0055] (2) Positive control

[0056] A positive control group (1% Vc ethyl ether) known to inhibit non-enzymatic glycosylation reactions was set up in each experiment.

[0057] (3) Blank control

[0058] The test substance and the positive control can emit a certain amount of fluorescence by themselves, so a blank control group is set up, that is, the test substance (or positive control) is mixed with PBS, and no reaction solution is added.

[0059] 2.4 Sample testing

[0060] (1) Prepare different test groups according to the ratios given in Table 1 and incubate in a 37°C constant temperature incubator away from light;

[0061] (2) On the 5th day, the fluorescence intensity was measured (excitation wavelength: 370 nm, emission wavelength: 440 nm).

[0062] Table 1 Reaction system (mL)

[0063] Trial Groups Test samples Positive PBS Reaction solution Sample set + - - + Negative control group - - + + Positive control group - + - + Blank control group + - + -

[0064] Note: In Table 1, “+” represents “added” and “-” represents “not added”.

[0065] 3. Test results

[0066] The calculation formula of inhibition rate (IR) is:

[0067]

[0068] RFU stands for Relative Fluorescence Unit. The test results are shown in Table 2.

[0069] Table 2 Non-enzymatic glycosylation inhibition results of each component

[0070]

[0071] The results showed that, as a single component, the inhibitory ability of sialic acid on non-enzymatic glycosylation was lower than that of bilberry extract and vitamin B 2 .

[0072] Test Example 2 Non-enzymatic glycosylation inhibition ability of each composition

[0073] Referring to the method of Experimental Example 1, the non-enzymatic glycosylation inhibition ability of the compositions of Examples 1-5 and Comparative Examples 1-2 was examined. The results are shown in Table 3.

[0074] Table 3 Non-enzymatic glycosylation inhibition results of each composition

[0075] Sample No. Test concentration (%) Inhibition rate (%) Example 1 5.0 87 Example 2 5.0 90 Example 3 5.0 86 Example 4 5.0 82 Example 5 5.0 90 Comparative Example 1 5.0 80 Comparative Example 2 5.0 89

[0076] The results show that although the compositions of Examples 1-5 do not use a single bilberry extract with a stronger inhibitory effect, the composition as a whole can effectively inhibit non-enzymatic glycosylation through the ratio of the components, and its inhibition level is not inferior to that of Comparative Example 2, and it is easier to make the final product components meet the concept of safe human intake in subsequent terminal applications.

[0077] Example 6

[0078] This embodiment provides a non-enzymatic glycosylation inhibitor in the form of tablets, and its raw material composition is as follows: 165 mg of sialic acid, 39 mg of ginkgo leaf extract, 90 mg of acerola extract, 6 mg of rosemary extract, 400 mg of microcrystalline cellulose, and 300 mg of sucrose powder.

[0079] Example 7

[0080] This embodiment provides a non-enzymatic glycosylation inhibitor in the form of an oral solution, and its raw material composition is as follows: 110 mg of sialic acid, 26 mg of ginkgo leaf extract, 60 mg of acerola extract, 4 mg of rosemary extract and 9800 mg of purified water.

[0081] Comparative Example 3

[0082] This comparative example provides a non-enzymatic glycosylation inhibitor, the dosage form is an oral solution, and the raw material composition is as follows: 98 mg of bilberry extract, 41 mg of ginkgo leaf extract, 48 mg of acerola extract, 4 mg of rosemary extract, and 4 mg of vitamin B 1 3mg, Vitamin B 2 3mg and vitamin B 6 3mg, purified water 9800mg.

[0083] Test Example 3 Stability test of each inhibitor

[0084] The oral liquids of Example 7 and Comparative Example 3 were sterilized by filtration with a 0.2 μm filter membrane, and then the sterilized samples were sealed in brown bottles and placed at 25°C for shelf life experiments. Sampling was performed at 30 days and 90 days to detect the sialic acid content and anthocyanin content. Each group of experiments was repeated 3 times, and the experimental results were the average of 3 times. The results are shown in Table 4.

[0085] Among them, the method for determining the sialic acid content is high performance liquid chromatography (HPLC), and the specific test conditions are as follows: Shimadzu Lc-15c; detection column Bio-Rad AMINEX HPX 87H Organic Analysis Column (300×7.8mm); column temperature 60°C; mobile phase 6mmol sulfuric acid, flow rate 0.6ml / min; detection wavelength 210nm.

[0086] The anthocyanin content was determined by high performance liquid chromatography (HPLC), and the specific test conditions were as follows: C18 column; column temperature 30°C; mobile phase was 4% phosphoric acid-acetonitrile solution with a volume ratio of 90:10, flow rate 0.8 ml / min; detection wavelength 520 nm.

[0087] Table 4 Stability test results of each inhibitor

[0088] Initial content 30 days content 90 days content Example 7 (Sialic Acid) 1.06％ 0.98％ 0.96％ Comparative Example 3 (anthocyanidins) 0.19％ 0.15％ 0.09％

[0089] It can be seen from the above results that the non-enzymatic glycosylation inhibitor of the present invention not only has a good anti-aging effect, but also has good stability.

[0090] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. An anti-non-enzymatic glycosylation composition, characterized in that: The invention is composed of the following raw materials in parts by weight: 40-70 parts of sialic acid, 10-20 parts of ginkgo leaf extract, 20-30 parts of acerola cherry extract and 0.1-10 parts of rosemary extract.

2. The anti-non-enzymatic glycosylation composition according to claim 1, characterized in that: The invention is composed of the following raw materials in parts by weight: 55 parts of sialic acid, 13 parts of ginkgo leaf extract, 30 parts of acerola cherry extract and 2 parts of rosemary extract.

3. The anti-non-enzymatic glycosylation composition according to any one of claims 1 or 2, characterized in that: The anti-non-enzymatic glycosylation composition is further added with 5 to 10 parts of natto extract.

4. Use of the anti-non-enzymatic glycosylation composition according to any one of claims 1 to 3 in the preparation of health food or cosmetics with anti-aging function.

5. A non-enzymatic glycosylation inhibitor, characterized in that The invention comprises the anti-non-enzymatic glycosylation composition according to any one of claims 1 to 3 and auxiliary materials acceptable in health food science.

6. The non-enzymatic glycosylation inhibitor according to claim 5, characterized in that The dosage form of the non-enzymatic glycosylation inhibitor is an oral preparation, including tablets, powders, and oral liquids.

7. The non-enzymatic glycosylation inhibitor according to claim 6, characterized in that The daily intake of sialic acid in the non-enzymatic glycosylation inhibitor is not less than 50 mg.

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