CN114948813A

CN114948813A - Anti-saccharification and anti-aging natural product composition, preparation method thereof and application thereof in oral beauty products

Info

Publication number: CN114948813A
Application number: CN202210608037.7A
Authority: CN
Inventors: 姜燕飞
Original assignee: Beijing Qingyan Bozhi Health Management Co ltd
Current assignee: Beijing Qingyan Bozhi Health Management Co ltd
Priority date: 2022-05-31
Filing date: 2022-05-31
Publication date: 2022-08-30
Anticipated expiration: 2042-05-31
Also published as: CN114948813B

Abstract

The invention discloses a preparation method of a natural product extract composition and application of the natural product extract composition in preparing anti-saccharification and anti-aging oral beauty products. Specifically, the invention discloses a water extract of broccoli seeds, a bamboo leaf flavone extract and epigallocatechin gallate, wherein the weight percentage of the water extract of broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate is 80%: 10%: 10% of the formulated composition. The biochemical test shows that the composition has the activity of inhibiting protein glycosylation and resisting aging, the inhibiting activity is obviously higher than that of a single extract, and the composition has very low cytotoxicity and can be used as an anti-saccharification and anti-aging component for preparing an oral beauty product. The invention also discloses the use of the composition derived from the natural product as an oral beauty component for preparing an anti-glycation and anti-aging oral liquid.

Description

Natural product composition for resisting saccharification and aging, preparation method thereof and application thereof in oral beauty product

The technical field is as follows:

the invention relates to a preparation method of an anti-saccharification and anti-aging natural product composition and application of the composition in preparing an oral beauty product, belonging to the field of food.

Background art:

brownlee first reported the presence of advanced glycation end products (AGEs) in vivo in biological macromolecules in 1984. AGEs refers to a class of complex glycosylation products produced by biological macromolecules (nucleic acids, proteins and lipids) through non-enzymatic mechanisms. Non-enzymatic glycosylation reactions refer primarily to a series of chemical reactions that occur between the terminal free carbonyl group of a reducing monosaccharide molecule and the free amino group in nucleic acids, proteins and lipid macromolecules (FIG. 1). Studies have found that the in vivo AGEs products can be divided into three major classes: non-crosslinked non-fluorescent product, non-crosslinked fluorescent product. More than 20 AGEs products have been identified in human blood and tissues.

AGEs are toxic molecules that potentially threaten human health. For example, glycosylation of protein molecules in vivo may result in changes in the secondary structure of the protein molecule, thereby affecting the function of the protein until it causes cell damage. In addition, collagen is an important tissue protein in the skin, and is gradually saccharified with the increase of age and high-sugar eating habits, so that the skin becomes hard and black, and finally, the skin is aged and the beauty is affected. Therefore, if the glycosylation level of biological macromolecules in vivo can be effectively inhibited, the composition can help to improve some disease states, delay aging and maintain the youthful state of the skin. Initial studies found that some natural products with antioxidant activity were effective in inhibiting AGEs formation, i.e., Anti-glycation (Anti-glycation) activity. With the progress of research, researchers have found that more different kinds of natural products can inhibit AGEs formation. Natural products with anti-glycation and anti-aging activities include polyphenols, polysaccharides, terpenoids, vitamins, alkaloids, polypeptides, etc., which inhibit the formation of AGEs by different mechanisms.

The natural product is an important source of anti-saccharification and anti-aging active products, and the natural product extract is rich in various natural products. The natural product extract obtained by early screening of the project group has obvious anti-saccharification and anti-aging activity. Considering that different types of natural products resist the formation of AGEs by different mechanisms, the combination of different natural product extracts and the combination of different AGEs inhibition mechanisms strives to find an extract composition with high anti-glycation and anti-aging activity.

The research history of natural plants on beauty can be traced back to thousands of years, and in recent years, oral liquid containing natural plant extracts is more and more popular among consumers. The oral beauty product containing the natural plant extract has obvious effect and high safety, and is popular with consumers.

The water extract of broccoli seeds contains sulforaphane, which is absorbed by human body and reacts with myrosinase to generate sulforaphane. Sulforaphane not only has strong anticancer activity, but also has strong oxidation resistance, and is one of the currently accepted natural products with cancer prevention, anticancer and beauty treatment effects. The bamboo leaf flavonoid extract is a plant flavonoid preparation newly developed in China, and has excellent biological effects of resisting free radicals, oxidation, aging, bacteria and viruses, protecting heart and cerebral vessels, preventing and treating senile degenerative diseases and the like. The product has rich material source, clear functional factors, good edible safety, high and stable preparation quality and fresh and sweet taste, and is widely used for health care products and oral beauty products. Epigallocatechin gallate is the main component of green tea polyphenols, and has antibacterial, antiviral, antioxidant, arteriosclerosis resisting, thrombosis resisting, blood vessel hyperplasia resisting, antiinflammatory and antitumor effects. The antioxidant activity is at least 100 times of that of vitamin C and 25 times of that of vitamin E, and can protect cells and DNA from damage, resist pollution, sunburn and smoking, and prevent and treat skin aging and wrinkle. So far, reports on the pharmacodynamic activity of water extract of broccoli seeds, bamboo leaf flavonoid extract, epigallocatechin gallate, single component or composition in the aspect of inhibiting AGEs formation are not found.

The invention content is as follows:

the invention aims to solve the first technical problem of providing a composition formed by mixing three natural extracts and a preparation method thereof. Is prepared from water extract OF BROCCOLI SEED (SEED OF Broccoli), bamboo leaf flavone extract (Phyllostachys MEYERI), and Epigallocatechin gallate (Epigallocatechin gallate), and its preparation method.

The second technical problem to be solved by the present invention is to provide the activity of the above composition. Researches show that compared with a single-component natural extract, the composition of the three extracts has obvious anti-saccharification and anti-aging activities. And such compositions have low cytotoxicity.

A third technical problem to be solved by the present invention is to provide the use of the above composition, a composition of natural extracts exhibiting good anti-glycation, anti-aging activity, thus using this composition as a cosmetic ingredient for the preparation of an oral cosmetic product, such as an oral liquid.

In order to solve the technical problem, the invention adopts the following technical scheme:

the first aspect of the invention relates to a composition prepared by mixing three extracted natural products, namely broccoli seed water extract, bamboo leaf flavone extract and epigallocatechin gallate according to a certain proportion, and the preparation method comprises the following steps:

mixing the water extract of the broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate according to a certain weight ratio, wherein the content of the water extract of the broccoli seeds is more than that of the bamboo leaf flavone extract and the epigallocatechin gallate, so that the weight ranges of the water extract of the broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate are determined as follows: bamboo leaf flavone extract: 60-90% of epigallocatechin gallate: 5-20%: 5-20%, weighing and mixing uniformly.

A further preferred process for the preparation of the composition is as follows: water extract of broccoli seeds: bamboo leaf flavone extract: epigallocatechin gallate ═ 80%: 10%: 10 percent. And weighing and uniformly mixing according to the optimized weight percentage.

The second aspect of the present invention relates to the above-mentioned natural product derived composition capable of reducing glycosylation of albumin. In a specific research, the anti-saccharification and anti-aging activities of three single extracts and the composition are compared, and the result shows that the anti-saccharification and anti-aging activities of the composition are remarkably superior to those of the single extracts, and the composition is suitable to be used as an anti-saccharification and anti-aging component for preparing corresponding oral beauty products.

The third aspect of the invention relates to the use of the composition derived from natural products in the preparation of an oral cosmetic product (such as an oral liquid) with anti-glycation and anti-aging activities. Also provides a specific formula of the oral liquid containing the composition from natural products as a cosmetic component and a preparation method thereof.

Description of the drawings:

FIG. 1: mechanism of formation of advanced glycation end products (AGEs) and damage to the skin.

FIG. 2: effects of different components on AGEs formation in vitro. The seven detection components comprise blank group, model group, aminoguanidine, water extract of broccoli seed, bamboo leaf flavone extract, epigallocatechin gallate and preferable extract composition.

FIG. 3: effect of different fractions on age formation in different tissues of mice. The six detection components comprise blank group, model group, water extract of broccoli seed, bamboo leaf flavone extract, epigallocatechin gallate and preferable extract composition.

FIG. 4: the anti-aging activity of the natural extract composition of the invention. Wherein A is a blank group; b is D-galactose induced senescence; c is UV-induced aging; d is aminoguanidine of a positive control group; e is the water extract group of broccoli seeds; f is bamboo leaf flavone extract; g is epigallocatechin gallate group; h is the preferred combination set.

The specific implementation mode is as follows:

the invention is further illustrated with reference to specific examples. It should be understood that the specific embodiments described herein are merely illustrative of the invention and do not limit the scope of the invention.

The examples do not show the specific techniques or conditions, according to the technical or conditions described in the literature in the field, or according to the product specifications. The reagents or instruments used are conventional products which are not indicated by manufacturers and are available from normal sources.

The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples are all commercially available products unless otherwise specified.

Example 1: preparation method of traditional Chinese medicine extract composition

Water extract OF BROCCOLI SEED (SEED OF BROCCOLI), off-white to brown powder, mild odor, available from BRASSICA corporation. Bamboo leaf flavone extract (Phyllostachys MEYERI), yellow or brown yellow powder, was purchased from Sheng's Biotech, Inc., Zhejiang. Epigallocatechin gallate (Epigallocatechin gate), an off-white to pale pink powder, loose and non-caking, purchased from St.Sci, Zhejiang Biotech Ltd. The preparation method of the composition comprises the following steps: the composition is prepared by mixing a broccoli seed water extract, a bamboo leaf flavone extract and epigallocatechin gallate according to a certain weight ratio, wherein the content of the broccoli seed water extract is more than that of the bamboo leaf flavone extract and the epigallocatechin gallate, and the weight ranges of the broccoli seed water extract, the bamboo leaf flavone extract and the epigallocatechin gallate are as follows: bamboo leaf flavone extract: epigallocatechin gallate 60-90%: 5-20%: 5-20%, weighing and mixing uniformly.

A further preferred method of preparation of the composition is as follows: according to the water extract of broccoli seeds: bamboo leaf flavone extract: epigallocatechin gallate 80%: 10%: weighing 10% of the mixture by weight and mixing the mixture evenly.

In the specific operation, 8g of broccoli seed water extract, 1g of bamboo leaf flavone extract and 1g of epigallocatechin gallate are weighed, and 10g of light yellow powder is mixed uniformly to obtain the composition disclosed by the invention.

Example 2: anti-glycation activity of natural extract composition

The experimental method comprises the following steps: the inhibitory activity of the water extract of broccoli seeds, the extract of bamboo leaf flavonoids, epigallocatechin gallate and the preferred composition prepared in example 1 on the formation of AGEs was examined. 0.5mL of the sample was added to 0.1M PBS buffer containing 0.5mL of 5.0g/mL BSA, 0.5mL of 0.5M glucose, and 1.0mL of 1% sodium azide at an aminoguanidine concentration of 4mg kg ^-1 Water extraction of broccoli seedsThe concentrations of extract, bamboo leaf flavone extract, epigallocatechin gallate and preferable composition are 0.03 mg/kg ^-1 . After incubation at 37 ℃ for 28 days, samples were taken on day 7, 14, 21 and 28, respectively, and measured by fluorescence spectrophotometry under conditions of Ex 370nm and Em 440nm, respectively, and Fluorescence Intensity (FI) of AGEs was measured. Wherein the blank group is a complete blank control, i.e. the generation of AGEs is not induced, and no inhibitor is added; no inhibitor was added to the model group. Anti-glycation reference positive compounds were compared using aminoguanidine.

The results are as follows:

table 1 effect of different components on AGEs formation: (

n＝4)

Note: 1) p in comparison with blank group<0.01; 2) in comparison with the model group, P<0.01; 3) the concentration of aminoguanidine group is 4 mmol.L ^-1 。

The results of the experiment are shown in table 1 and fig. 2. Lower AGEs fluorescence values indicate better anti-glycation. Aminoguanidine, water extract of broccoli seed, bamboo leaf flavone extract, epigallocatechin gallate and preferable composition in different days can inhibit AGEs generation, and have certain anti-saccharification activity. Data analysis was performed using Graphpad Prism 9.3.1, and using multiple comparisons in two-way anova, it was preferred whether the compositions were statistically different from each other group in inhibition of AGEs formation on different days. The results demonstrate that the inhibitory activity of the preferred compositions on AGEs formation is significantly superior to the aminoguanidine group and each single component group and is statistically significant (P < 0.0001). The water extract of the broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate are proved to be 80 percent: 10%: the preferred composition resulting from the 10% weight percentage blend has significant anti-glycation activity and is superior to aminoguanidine.

Example 3: effect of Natural extract composition on AGE production in diabetic mouse tissues

The experimental method comprises the following steps: 50 Balb/c mice (SPF grade), male and female, were purchased from Schbefu, Beijing Biotechnology Ltd. A diabetic mouse model is established to study the influence of different natural isolates on AGEs in organs such as heart, liver and kidney. Intravenous injection of tetraoxypyrimidine (80mg kg) at mouse tail ^-1 ) And establishing a diabetic mouse model. 50 eligible mice were selected and randomly divided into model and experimental groups (three natural product extracts and preferred compositions), 10 per group. And taking 10 healthy mice as a blank group, wherein the blank group does not construct a diabetes mouse model and does not perform gastric lavage. The model group and the experimental group are respectively irrigated with physiological saline, three natural product extracts and a preferable composition, the dosage of the irrigated stomach is 200 mu L/mouse, and the concentration of the irrigated stomach of each group is 30mg kg ^-1 Once daily for 14 days. Mice were fasted for 16 hours after the last dose, and then sacrificed by cervical dislocation for follow-up experiments. Mouse heart, liver and kidney tissues were cut into small pieces, homogenized and centrifuged. 5mL of chloroform-methanol (2:1) was added to the precipitate for defatting and shaking at 4 ℃ overnight. Then, after washing in HEPES buffer and overnight, collagenase was added and digested at 37 ℃ for 24 hours. The above solution was centrifuged to obtain a supernatant, and the fluorescence intensity was measured at 370nm/440nm using a fluorescence spectrophotometer. Relative fluorescence intensity of the samples was obtained according to HEPES buffer.

The results are as follows:

TABLE 2 Effect of different fractions on AGEs formation in different tissues of mice: (

n＝10)

Note: 1) p <0.01 compared to blank; 2) p <0.01 compared to model group.

The results of the experiment are shown in table 2 and fig. 3. Lower AGEs fluorescence values indicate better anti-glycation. The water extract of broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate and the preferable composition inhibit the AGEs in various organs of the diabetic mouse to different degrees in vivo, and the inhibition rate of the AGEs generated by the heart and the liver is better than that of the kidney. Data analysis was performed using Graphpad Prism 9.3.1, and multiple comparisons in a two-way anova were used to compare whether the inhibition of AGEs formation was statistically different in different tissues of mice, preferably between the composition and each other group. The results show that the inhibition rate of the preferred composition on AGEs formation in the diabetic mouse tissue is significantly better than that of each single component group, and has statistical significance (P <0.0001), which proves that the water extract of broccoli seeds, the bamboo leaf flavonoid extract and the epigallocatechin gallate are mixed according to the ratio of 80%: 10%: the preferred composition resulting from the 10% weight percentage blend has significant in vivo anti-glycation activity.

Example 4: anti-aging activity of natural extract composition

The experimental method comprises the following steps: the detection of cellular senescence was observed by staining for senescence-associated beta-galactosidase (SA-beta-gal). Cell senescence beta-galactosidase staining kit was used for detection. Specifically, CP-H103 cells were seeded into 6-well plates at approximately 50000 cells/2 mL culture per well. CP-H103 cells were stimulated with 400. mu.g/mL aminoguanidine, 100. mu.g/mL broccoli seed water extract, 100. mu.g/mL bamboo leaf flavonoid extract, 100. mu.g/mL epigallocatechin gallate and 100. mu.g/mL extract combination, UV radiation (222mM) and D-galactose, respectively, to simulate sun aging and skin aging. After passage 5 times, the cell culture fluid was aspirated, washed 1 time with PBS, 1mL of β -galactosidase staining fixative was added, fixed for 15 minutes at room temperature, the cell fixative was aspirated, and the cells were washed 3 times with PBS, 3 minutes each. PBS was aspirated and 1mL of staining solution was added to each well. Incubate overnight at 37 ℃ and seal 6-well plates with preservative film to prevent evaporation. Observed under a normal optical microscope.

Observation indexes are as follows: senescent cells produce dark blue products under the catalysis of beta-galactosidase. Thus, the beta-galactosidase expressing cells or tissues changed to blue color were easily observed under an optical microscope.

The results are as follows:

as can be seen from fig. 4, the phenomena of D-galactose-induced aging and UV-induced aging of sun-aged cells are significant, although the broccoli seed water extract group, the bamboo leaf flavonoid extract, and the epigallocatechin gallate group all have a certain inhibitory effect on aging of cells, the anti-aging effect is lower than that of aminoguanidine in the positive control group, and the anti-aging effect of the preferred composition of the three natural extract compositions is significantly better than that of aminoguanidine in the positive control group, thereby proving that the preferred composition has a good anti-aging activity.

Example 5: cytotoxicity of Natural extract composition

The experimental method comprises the following steps: the cytotoxicity of the compound is detected by adopting a Beyotime kit. The specific operation is to respectively inoculate HepG2 and 293T, A549 cells into a 96-well plate and culture for 6-8 hours. After treating with 400. mu.g/mL aminoguanidine, 100. mu.g/mL broccoli seed water extract, 100. mu.g/mL bamboo leaf flavone extract, 100. mu.g/mL epigallocatechin gallate and 100. mu.g/mL preferred composition for 24 hours, the cell activity was analyzed by MTT method. The absorbance was then measured at 570nm with a microplate reader.

The results are as follows:

TABLE 3 cytotoxicity of different components on mammalian cells

The results are shown in table 3, and the cell experiment tests show that the broccoli seed water extract, the bamboo leaf flavone extract, the epigallocatechin gallate and the preferable composition of the three have no obvious cytotoxicity. The combination of example 2, example 3, the three, especially preferred, is suitable for use as an anti-glycation, anti-ageing ingredient in the preparation of a cosmetic product.

Example 6: natural extract composition used as oral beauty component for preparing anti-saccharification and anti-aging oral liquid

An anti-saccharification and anti-aging oral liquid comprises the following formula and parts by weight:

68.79 parts of drinking water, 10 parts of red grape concentrated juice, 10 parts of orange concentrated juice, 5 parts of erythritol, 5 parts of sweet orange powder, 0.8g of broccoli seed water extract, 0.1 part of bamboo leaf flavone, 0.1 part of epigallocatechin gallate, 0.1 part of citric acid, 0.1 part of sodium citrate and 0.01 part of edible essence.

The preparation method comprises the following steps: according to the above formulation, the respective components were weighed, wherein each mass part was 1 g. Mixing and stirring uniformly, placing into 300mL glass bottle, boiling at 85 deg.C for 20 min for sterilization, cooling to 25 deg.C, and bottling.

It will be apparent to those skilled in the art that various changes and modifications may be made in the present invention without departing from the spirit and scope of the invention. Thus, if such modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include such modifications and variations.

Claims

1. A composition of natural product origin, characterized in that: comprises the following components in percentage by weight: water extract of broccoli seeds: bamboo leaf flavone extract: 60-90% of epigallocatechin gallate: 5-20%: 5 to 20 percent.

2. The natural product derived composition according to claim 1, characterized in that: comprises the following components in percentage by weight: water extract of broccoli seeds: bamboo leaf flavone extract: epigallocatechin gallate 80%: 10%: 10 percent.

3. A method of preparing a natural product derived composition according to claim 1 or claim 2, characterized in that: the composition of natural product source is prepared by weighing the water extract of broccoli seeds, the bamboo leaf flavone extract and the epigallocatechin gallate according to the weight percentage in claim 1 or claim 2, and mixing the three powders.

4. Use of a composition of natural product origin according to claim 1 or claim 2, for the preparation of an oral cosmetic product, characterized in that: the oral cosmetic product has anti-glycation activity for inhibiting formation of advanced glycation end products.

5. Use of a composition of natural product origin according to claim 1 or claim 2, for the preparation of an oral cosmetic product, characterized in that: the oral cosmetic product has anti-aging activity.

6. Use according to claim 4 or claim 5, characterized in that: the beauty treatment oral product is oral liquid.

7. An anti-saccharification and anti-aging oral liquid is characterized in that: the oral liquid contains the composition derived from the natural product as described in claim 1 or claim 2, and the formula and the parts by weight of the oral liquid are as follows: 5 parts of the natural product derived composition of claim 1 or claim 2, 68.79 parts of drinking water, 10 parts of red grape juice concentrate, 10 parts of orange juice concentrate, 5 parts of erythritol, 5 parts of sweet orange powder, 0.1 part of citric acid, 0.1 part of sodium citrate, 0.01 part of flavoring essence.