Anti-sugar and anti-oxidation composition as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of food, and particularly relates to an anti-sugar and anti-oxidation composition as well as a preparation method and application thereof. A mulberry leaf extract, an emblic leafflower fruit extract, pleurotus citrinopileatus powder and rosa roxburghii tratt fruit powder are used as composite plant fermentation raw materials, Weissella helenii and pediococcus pentosaceus are added for mixed fermentation, a composite plant fermentation product is prepared, then lecithin and vitamin E are added, and the anti-sugar and anti-oxidation composition is obtained. The composition has excellent anti-sugar and anti-oxidation effects, and is safe and non-irritant. The preparation method of the anti-sugar and anti-oxidation composition is simple, can be applied to preparation of anti-sugar and anti-oxidation products, and has good market application value.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of food technology, and particularly relates to an anti-sugar and anti-oxidation composition, a preparation method and application thereof. BACKGROUND

[0002] Skin aging is caused by the combined action of multiple factors. Non-enzymatic glycosylation (NEG) refers to a process in which biological macromolecules such as proteins, lipids, nucleic acids, etc. spontaneously react with glucose or other reducing monosaccharides under non-enzymatic conditions to produce a substance called advanced glycation end products (AGEs). The formation of AGEs is a complex, irreversible, multi-step process, in which carboxymethyl lysine (CML) and pentoside are the most common AGEs in the skin. With age, AGEs accumulate in the body, causing cross-linking of adjacent proteins, carbohydrates, lipids, nucleic acids and other substances, not only affecting the structure of these substances, causing changes in their biological properties, leading to loss of elasticity, reducing the skin's ability to scavenge free radicals, and weakening the skin's antioxidant properties. AGEs also make the skin lack moisture, yellow, loose, and increase wrinkles, thereby accelerating skin aging. The so-called "anti-glycation" is actually anti-AGEs. Oxidative stress is a negative effect caused by free radicals in the body and is considered an important factor in causing aging and diseases. Persistent oxidative stress can lead to aging, cancer, cardiovascular disease and Alzheimer's disease.

[0003] For anti-glycation, although many synthetic and natural ingredients have been proven to have AGEs generation inhibiting effect, there is still a long way to go to effectively resist glycation. For example, most polyphenols have strong antioxidant and AGEs inhibiting activity, but polyphenols are unstable, easily oxidized and inactivated, and have low bioavailability, making it difficult to play a role in the body. For anti-oxidation, artificial antioxidants are commonly used in the prior art, which can directly react with superoxide anion free radicals, hydroxyl free radicals, hydrogen peroxide and other free radicals, and remove them by providing electrons or hydrogen atoms, thereby reducing the oxidative damage of free radicals to skin cells. However, common artificial antioxidants such as tert-butyl hydroquinone, dibutyl hydroxytoluene and butylated hydroxyanisole have side effects such as affecting body metabolism, causing allergic reactions and potential carcinogenicity in the application process.

[0004] In summary, there is an urgent need to develop a product with anti-sugar and anti-oxidation effects, and safety, no irritation and no side effects, to solve the problems of poor anti-glycation effect, strong irritation and poor safety of existing anti-sugar and anti-oxidation products. SUMMARY

[0005] In view of the above-mentioned defects or improvement needs of the prior art, the present invention provides an anti-glycation and antioxidant composition, its preparation method and application, so as to solve the problems of poor anti-glycation effect, strong irritation and poor safety of existing anti-glycation and antioxidant products.

[0006] The objective of this invention can be achieved through the following technical solutions:

[0007] A first aspect of the present invention is to provide an anti-glycation and antioxidant composition comprising the following components: a complex plant fermentation product, lecithin, and vitamin E.

[0008] The compound plant fermentation product is obtained by fermenting mulberry leaf extract, amla extract, pine nut powder and prickly pear powder as compound plant fermentation raw materials, and inoculating them with compound bacteria.

[0009] Mulberry leaf (Folium Mori) is the dried leaf of the mulberry tree, a plant in the Moraceae family, including white mulberry (Morus Alba), red mulberry (Morus Rubra), and black mulberry (Morus Nigra). Mulberry leaf extract (MLE) is rich in flavonoids, alkaloids, polysaccharides, and other bioactive substances, and has antioxidant, anti-inflammatory, glucose and lipid metabolism regulating, and immune-enhancing effects.

[0010] Phyllanthus emblica is a perennial deciduous small tree or shrub belonging to the genus Phyllanthus in the family Euphorbiaceae. It is mainly distributed in tropical and subtropical regions of South Asia. Phyllanthus emblica extract contains a variety of active ingredients such as polyphenols, flavonoids, polysaccharides, organic acids, and terpenes, as well as inorganic elements such as potassium, magnesium, iron, and zinc. It is a natural antioxidant and has anti-inflammatory, antibacterial, anti-tumor, lipid-lowering, anti-aging, and cardiovascular-enhancing effects.

[0011] Golden-topped oyster mushroom, also known as golden-topped mushroom, elm yellow mushroom, or jade emperor mushroom, contains bioactive components such as fumaric acid, niacin, D-mannitol, ergosterol, and lovastatin. It has the effects of enhancing immunity, anti-oxidation, anti-tumor, antibacterial, promoting cell metabolism, and lowering blood lipids.

[0012] Prickly pear is the fruit of *Rosa latifolia* and *Rosa multiflora*, plants belonging to the genus *Rosa* of the Rosaceae family. It has a sweet and sour taste and is effective in strengthening the stomach, aiding digestion, and stopping diarrhea. It contains active substances such as polyphenols and flavonoids, and has antioxidant, antibacterial, antitumor, anti-inflammatory, immune-enhancing, and lipid-regulating effects.

[0013] Preferably, the compound bacteria consists of *Helenweissella* and *Pediococcus pentosus* in a volume ratio of 1:1-2.

[0014] More preferably, the compound bacteria consists of *Helenweissella* and *Pediococcus pentosus* in a volume ratio of 1:1.3.

[0015] Preferably, the preservation number of the *Helenweissella* is CGMCC NO.1.2513, and the preservation number of the *Pediococcus pentosaceus* is CGMCC NO.1.7665.

[0016] Preferably, the viable count of the *Herrenweissella* is 3-5 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 3-5 × 10⁻⁶ CFU / mL. 8 CFU / mL.

[0017] More preferably, the viable count of the *Herrenweissella* is 4 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 4 × 10⁻⁶. 8 CFU / mL.

[0018] Preferably, the compound plant fermentation raw material includes the following raw materials in weight percentage: 8%-12% mulberry leaf extract, 10%-15% amla extract, 5%-8% pine nut powder, and 6%-10% prickly pear powder.

[0019] More preferably, the compound plant fermentation product is prepared from the following raw materials in weight percentage: 10% mulberry leaf extract, 13% amla extract, 7% pine nut powder, 8% prickly pear powder and the remainder being deionized water.

[0020] Preferably, the amount of the compound bacteria inoculated is 3-5% of the weight of the compound plant fermentation raw materials.

[0021] A second aspect of this invention aims to provide a method for preparing the composition described in the first aspect of this invention, specifically comprising the following steps:

[0022] A1. Weigh each component of the compound plant fermentation raw material described in the first aspect according to the formula, add deionized water and mix evenly to obtain the fermentation substrate;

[0023] A2. Inoculate the activated compound bacterial solution into the fermentation substrate for fermentation. After fermentation, filter and concentrate the solution, then add lecithin and vitamin E, and homogenize to obtain the fermentation composition.

[0024] Preferably, the fermentation temperature in step A2 is 30-35℃ and the fermentation time is 24-48h.

[0025] Preferably, the amount of lecithin added in step A2 is 0.2-0.4% of the total weight of the compound plant fermentation raw materials.

[0026] Preferably, the amount of vitamin E added in step A2 is 0.06%-0.08% of the total weight of the compound plant fermentation raw materials.

[0027] The object of a third aspect of the present invention is to provide the use of the composition described in the first aspect of the present invention or the composition prepared by the preparation method described in the second aspect of the present invention in the preparation of anti-glycation and antioxidant products.

[0028] Preferably, the dosage form of the product includes tablets, powders, pills, liquids, or capsules.

[0029] The beneficial effects of this invention are:

[0030] This invention provides an anti-glycation and antioxidant composition. It is prepared by fermenting a compound plant product using mulberry leaf extract, amla extract, pleurum root powder, and prickly pear powder as fermentation raw materials, adding a compound microbial culture. Lecithin and vitamin E are then added and homogenized to obtain the anti-glycation and antioxidant composition. Mulberry leaf extract has antioxidant, anti-inflammatory, glucose and lipid metabolism regulating, and immune-enhancing effects; amla extract has anti-inflammatory, antibacterial, anti-tumor, lipid-lowering, anti-aging, and cardiovascular-enhancing effects; pleurum root powder has antioxidant, blood sugar-lowering, anti-inflammatory, cardiovascular-protective, and anti-cancer effects; and prickly pear powder has antioxidant, anti-inflammatory, vasodilatory, anti-allergic, and anti-diabetic effects. In this invention, the combination of mulberry leaf extract, amla extract, pleurum root powder, and prickly pear powder synergistically enhances the anti-glycation and antioxidant effects of the fermentation product, thereby improving the efficacy of the composition containing this fermentation product. The *Vibrio hurenweiss* selected in this invention belongs to heterofermentative lactic acid bacteria. It not only produces antibacterial substances such as bacteriocins and organic acids, but also extracellular polysaccharides. Its fermentation products have anti-inflammatory, antioxidant, cholesterol-lowering, antibacterial, and oral health-improving effects. *Pediococcus pentosaceus*, a probiotic belonging to the genus *Pediococcus* of the family Lactobacillus, belongs to homofermentative lactic acid bacteria. It enhances host immunity, improves oxidative stress levels, maintains intestinal flora balance, and regulates the body's immune response, thus having a positive effect on human health. Experimental data provided by this invention show that when *Vibrio hurenweiss* and *Pediococcus pentosaceus* are fermented in a specific ratio, the resulting composition exhibits superior antioxidant and antiglycation effects. This indicates that the mixed fermentation of *Pediococcus pentosaceus* (homogeneous lactic acid bacteria) and *Vibrio hurenweiss* (heterogeneous lactic acid bacteria) allows for metabolic complementarity and synergistic regulation, which is beneficial for improving fermentation efficiency, increasing the yield and stability of active substances.

[0031] The preparation method of the anti-glycation and antioxidant composition of the present invention is simple. By fermenting the compound plant fermentation raw materials with a mixed culture of Pediococcus pentosaceus and Helenweissella, the anti-glycation and antioxidant effects and stability of the active ingredients in the composition can be effectively improved. The prepared composition has excellent anti-glycation and antioxidant effects and can be used to prepare anti-glycation and antioxidant products. It has good market application value and can solve the problems of poor anti-glycation effect, strong irritation and poor safety of existing anti-glycation and antioxidant products. Detailed Implementation

[0032] To further illustrate the technical means and effects of the present invention in achieving its intended purpose, the following detailed description of the specific implementation methods, structures, features and effects of the present invention, in conjunction with embodiments, is provided below.

[0033] Unless otherwise specified, experimental methods in the following examples are generally performed under standard conditions or as recommended by the manufacturer. All materials and reagents used are commercially available unless otherwise specified.

[0034] Mulberry leaf extract was purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd.;

[0035] Phyllanthus emblica extract was purchased from Nanjing Haolan Health Technology Co., Ltd.;

[0036] The Golden Top Orchid Powder was purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd.

[0037] The prickly pear powder was purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd.

[0038] Compound plant fermentation raw materials ①

[0039] The compound plant fermentation raw material is prepared from the following raw materials in weight percentage: 10% mulberry leaf extract, 13% amla extract, 7% pine nut powder, 8% prickly pear powder and the remainder deionized water.

[0040] Compound plant fermentation raw materials ②

[0041] The compound plant fermentation raw material is prepared from the following raw materials in weight percentage: 8% mulberry leaf extract, 10% amla extract, 5% pine nut powder, 6% prickly pear powder and the remainder deionized water.

[0042] Compound plant fermentation raw materials ③

[0043] The compound plant fermentation raw material is prepared from the following raw materials in weight percentage: 12% mulberry leaf extract, 15% amla extract, 8% golden top pine ear powder, 10% prickly pear powder and deionized water as the balance.

[0044] Compound plant fermentation raw materials ④

[0045] The only difference between compound plant fermentation raw material ④ and compound plant fermentation raw material ① is that compound plant fermentation raw material ④ does not contain mulberry leaf extract and amla extract, and the reduced weight of these is allocated to Pleurotus ostreatus powder and Prickly pear powder in a 7:8 ratio.

[0046] Compound plant fermentation raw materials ⑤

[0047] The only difference between compound plant fermentation raw material ⑤ and compound plant fermentation raw material ① is that compound plant fermentation raw material ⑤ does not contain Pleurotus ostreatus powder, and its reduced weight is allocated to mulberry leaf extract, amla extract and prickly pear powder in a ratio of 10:13:8.

[0048] Compound plant fermentation raw materials⑥

[0049] The only difference between compound plant fermentation raw material ⑥ and compound plant fermentation raw material ① is that: no prickly pear powder is added to compound plant fermentation raw material ⑥, and its reduced weight is allocated to mulberry leaf extract, amla extract and golden top pine mushroom powder in a ratio of 10:13:7.

[0050] Example 1

[0051] A method for preparing an anti-glycation and antioxidant composition specifically includes the following steps:

[0052] A1. Weigh the compound plant fermentation raw material ① according to the formula, mix them evenly, and obtain the fermentation substrate;

[0053] A2. Inoculate the activated compound bacterial solution into the fermentation substrate and ferment at a temperature of 33°C for 36 hours. After fermentation, filter and concentrate the solution, add lecithin and vitamin E, and homogenize to obtain the fermentation composition.

[0054] In step A2, the total amount of compound bacteria added is 4% of the weight of the compound plant fermentation raw materials;

[0055] In step A2, the compound bacteria consist of *Helicobacter schrenckii* and *Pediococcus pentosus* in a volume ratio of 1:1.3. The preservation number of *Helicobacter schrenckii* is CGMCC NO.1.2513, and the preservation number of *Pediococcus pentosus* is CGMCC NO.1.7665.

[0056] The viable count of *Helenweissella* in step A2 is 4 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 4 × 10⁻⁶. 8 CFU / mL.

[0057] In step A2, the amount of lecithin added is 0.3% of the total weight of the compound plant fermentation raw materials.

[0058] In step A2, the amount of vitamin E added is 0.07% of the total weight of the compound plant fermentation raw materials.

[0059] Example 2

[0060] A method for preparing an anti-glycation and antioxidant composition specifically includes the following steps:

[0061] A1. Weigh the compound plant fermentation raw material ① according to the formula, mix them evenly, and obtain the fermentation substrate;

[0062] A2. Inoculate the activated compound bacterial solution into the fermentation substrate and ferment at a temperature of 30°C for 48 hours. After fermentation, filter and concentrate the solution, add lecithin and vitamin E, and homogenize to obtain the fermentation composition.

[0063] In step A2, the total amount of compound bacteria added is 3% of the weight of the compound plant fermentation raw materials;

[0064] In step A2, the compound bacteria consist of *Helicobacter schrenckii* and *Pediococcus pentosus* in a volume ratio of 1:1. The preservation number of *Helicobacter schrenckii* is CGMCC NO.1.2513, and the preservation number of *Pediococcus pentosus* is CGMCC NO.1.7665.

[0065] The viable count of *Helenweissella* in step A2 is 3 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 5 × 10⁻⁶. 8 CFU / mL.

[0066] In step A2, the amount of lecithin added is 0.2% of the total weight of the compound plant fermentation raw materials.

[0067] In step A2, the amount of vitamin E added is 0.06% of the total weight of the compound plant fermentation raw materials.

[0068] Example 3

[0069] A method for preparing an anti-glycation and antioxidant composition specifically includes the following steps:

[0070] A1. Weigh the compound plant fermentation raw material ① according to the formula, mix them evenly, and obtain the fermentation substrate;

[0071] A2. Inoculate the activated compound bacterial solution into the fermentation substrate and ferment at a temperature of 35°C for 24 hours. After fermentation, filter and concentrate the solution, add lecithin and vitamin E, and homogenize to obtain the fermentation composition.

[0072] In step A2, the total amount of compound bacteria added is 5% of the weight of the compound plant fermentation raw materials;

[0073] In step A2, the compound bacteria consist of *Helicobacter schrenckii* and *Pediococcus pentosus* in a volume ratio of 1:2. The preservation number of *Helicobacter schrenckii* is CGMCC NO.1.2513, and the preservation number of *Pediococcus pentosus* is CGMCC NO.1.7665.

[0074] In step A2, the viable count of *Helenweissella* is 5 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 3 × 10⁻⁶. 8 CFU / mL.

[0075] In step A2, the amount of lecithin added is 0.4% of the total weight of the compound plant fermentation raw materials.

[0076] In step A2, the amount of vitamin E added is 0.08% of the total weight of the compound plant fermentation raw materials.

[0077] Example 4

[0078] The only difference between Example 4 and Example 1 is that Example 4 uses compound plant fermentation raw material ②, while all other conditions are the same as in Example 1.

[0079] Example 5

[0080] The only difference between Example 5 and Example 1 is that Example 5 uses compound plant fermentation raw material ③, while all other conditions are the same as in Example 1.

[0081] Comparative Example 1

[0082] The only difference between Comparative Example 1 and Example 1 is that Comparative Example 1 uses compound plant fermentation raw material ④, while all other conditions are the same as in Example 1.

[0083] Comparative Example 2

[0084] The only difference between Comparative Example 2 and Example 1 is that Comparative Example 2 uses a compound plant fermentation raw material ⑤, while all other conditions are the same as in Example 1.

[0085] Comparative Example 3

[0086] The only difference between Comparative Example 3 and Example 1 is that Comparative Example 3 uses a compound plant fermentation raw material ⑥, while all other conditions are the same as in Example 1.

[0087] Comparative Example 4

[0088] The only difference between Comparative Example 4 and Example 1 is that in step A2 of Comparative Example 4, the fermentation substrate is inoculated with Pediococcus pentosacchari culture for fermentation (i.e., without adding Helenweiss bacteria), and the volume reduced by Helenweiss bacteria is made up with Pediococcus pentosacchari. All other conditions are the same as in Example 1.

[0089] Comparative Example 5

[0090] The only difference between Comparative Example 5 and Example 1 is that in step A2 of Comparative Example 5, the fermentation substrate is inoculated with Helenweiss bacteria (i.e., without adding Pediococcus pentosus), and the volume lost by Pediococcus pentosus is made up with Helenweiss bacteria. All other conditions are the same as in Example 1.

[0091] Comparative Example 6

[0092] The only difference between Comparative Example 6 and Example 1 is that the volume ratio of Helenweissella to Pediococcus pentosus in Comparative Example 6 is 1:0.5, and the total number of viable bacteria inoculated into the fermentation substrate and other conditions are the same as in Example 1.

[0093] Comparative Example 7

[0094] The only difference between Comparative Example 7 and Example 1 is that the volume ratio of Helenweissella to Pediococcus pentosus in Comparative Example 7 is 1:3, and the total number of viable bacteria inoculated into the fermentation substrate and other conditions are the same as in Example 1.

[0095] Example 1: In vitro inhibition of non-enzymatic glycosylation assay

[0096] By establishing a bovine serum albumin-glucose glycosylation reaction system to simulate non-enzymatic glycosylation reactions in the human body, the inhibitory performance of the composition on non-enzymatic glycosylation was tested.

[0097] Test substances: Compositions prepared in Examples 1-5 and Comparative Examples 1-7 of this invention;

[0098] The specific testing methods are as follows:

[0099] T1. The test substance was diluted with PBS buffer to a concentration of 1 wt% for later use.

[0100] T2. Using PBS buffer solution containing 1% NaN3, with a concentration of 0.20 mol / L and a pH of 7.4 as the diluent, take 10 mL each of 15 mg / mL bovine serum albumin solution and 0.3 mol / L glucose solution, mix them evenly, and use them as the reaction solution.

[0101] T3. Add 10 mL of the test substance to the reaction solution and incubate at 37°C in the dark for 14 days. For the positive control group, use 10 mL of 1 wt% aminoguanidine sulfate instead of the test substance. For the negative control group, use 10 mL of PBS buffer solution containing 1% NaN3, with a concentration of 0.20 mol / L and a pH of 7.4 instead of the test substance. For the blank control group, use 20 mL of PBS buffer solution containing 1% NaN3, with a concentration of 0.20 mol / L and a pH of 7.4 instead of the reaction solution and test substance. Three parallel tubes are used for each group.

[0102] T4: On day 14, the fluorescence value (RFF) of each group at the excitation wavelength of 370 nm and the emission wavelength of 440 nm was measured using a fluorescence microplate reader. The inhibition rate of non-enzymatic glycosylation was calculated according to the following formula:

[0103]

[0104] Among them, RFU 受试物组 The fluorescence value of the test group;

[0105] RFU 空白对照组 The fluorescence value is for the blank control group;

[0106] RFU 阴性对照组 The fluorescence value is for the negative control group;

[0107] The test results are shown in Table 1.

[0108] Table 1. Data on non-enzymatic glycosylation inhibition rate

[0109]

[0110]

[0111] Example 2: Antioxidant Performance Test

[0112] DPPH (1,1-diphenyl-2-trinitrophenylhydrazine) is a stable free radical. The DPPH scavenging rate of the compositions prepared in Examples 1-5 and Comparative Examples 1-7 was determined, and the specific operating procedures are as follows:

[0113] S1. The test sample was prepared into a 0.5 wt% test solution with deionized water and a 0.2 mmol / L DPPH free radical solution was prepared with anhydrous ethanol.

[0114] S2. For each sample group, take 2 mL of the test solution and place it in a test tube, then add 2 mL of the previously prepared DPPH free radical solution; take 2 mL of deionized water and place it in a test tube, then add 2 mL of the previously prepared DPPH free radical solution as the control group; take 2 mL of the test solution and place it in a test tube, then add 2 mL of anhydrous ethanol as the blank group; after mixing the above groups, react at room temperature in the dark for 30 min, then measure the absorbance value at 517 nm. Each mixture should be measured in triplicate, and the scavenging rate should be calculated using the following formula:

[0115]

[0116] The test results are shown in Table 2.

[0117] Table 2 DPPH free radical scavenging rate data

[0118]

[0119] Compared with Example 1, Comparative Examples 1-3 did not add mulberry leaf extract and amla extract, golden pine nut powder and prickly pear powder, respectively; Comparative Examples 4-5 had different fermentation strains, respectively inoculated with only Pediococcus pentosus or Helenweissella for single-strain fermentation; and Comparative Examples 6-7 had different strain ratios and were not within the range of the ratios of the present invention.

[0120] Based on the data from Examples 1 and 4-5 in Tables 1-2, the non-enzymatic glycosylation inhibition rate and DPPH free radical scavenging rate of Examples 4-5 decreased, indicating that the compound plant fermentation raw materials prepared from 10% mulberry leaf extract, 13% amla extract, 7% pine nut powder and 8% prickly pear powder have the best synergistic effect, thereby improving the anti-glycation and antioxidant efficacy of the composition.

[0121] Based on the data from Example 1 and Comparative Examples 1-3 in Table 1-2, the non-enzymatic glycosylation inhibition rate and DPPH free radical scavenging rate of Comparative Examples 1-3 were significantly reduced, indicating that the combination of mulberry leaf extract, amla extract, pine nut powder, and prickly pear powder synergistically enhanced the anti-glycation and antioxidant effects of the composition. Based on the data from Example 1 and Comparative Examples 4-7 in Table 1-2, the non-enzymatic glycosylation inhibition rate and DPPH free radical scavenging rate of Comparative Examples 4-7 were also significantly reduced, indicating that the composition and ratio of the strains both affect the anti-glycation and antioxidant effects of the composition. Furthermore, based on the data from Example 1 and Comparative Examples 4-5 in Table 1-2, it is shown that the mixed fermentation of Pediococcus pentosaceus and Helenweissella has a synergistic effect, which is beneficial to improving fermentation efficiency, increasing the yield and stability of active substances, thereby enhancing the anti-glycation and antioxidant efficacy of the composition.

[0122] Based on the data from Comparative Examples 4-5 and 6-7 in Table 1-2, the non-enzymatic glycosylation inhibition rate and DPPH free radical scavenging rate of Comparative Example 4-5 were lower than those of Comparative Example 6-7, indicating that the composition of the fermentation strains has a greater impact on the anti-glycation and antioxidant effects of the composition than the strain ratio. The optimal volume ratio of Helenweissella to Pediococcus pentosus is 1:1.3.

[0123] In summary, this invention uses a compound of mulberry leaf extract, amla extract, pleurum root powder, and prickly pear powder, which synergistically enhances anti-glycation and antioxidant effects. By using a mixed-culture fermentation process of Pediococcus pentosaceus and Helenweissella var. humilis to ferment the compound plant fermentation raw materials, the anti-glycation and antioxidant effects and stability of the active ingredients in the composition can be effectively improved. The prepared composition has excellent anti-glycation and antioxidant efficacy, which can solve the problems of poor anti-glycation effect, strong irritation, and poor safety of existing anti-glycation and antioxidant products.

[0124] Example 3: Cytotoxicity Test

[0125] Sample: The compositions prepared in Examples 1-5 of this invention;

[0126] The specific experimental steps are as follows:

[0127] A1. Take stable RAW264.7 cells, resuspend them in DMEM complete medium, and adjust the cell concentration to 2×10⁻⁶. 5 Cells / mL, 100 μL per well was seeded into 96-well cell culture plates (except for the outermost wells and blank control wells), and cultured at 37℃, 5% CO2 and saturated humidity for 24 h;

[0128] A2. Remove the culture medium, add 100 μL of sample solution diluted with PBS to a concentration of 100 μg / mL to each well according to the concentration gradient, and add an equal amount of PBS solution to the control wells. Set up 6 replicates for each group. After incubation for 24 h, remove the solution and wash twice with PBS.

[0129] A3. Add 100 μL of fresh DMEM medium to each well, then add 10 μL of MTT reagent, and transfer the 96-well plate to an incubator to continue incubation for 4 hours.

[0130] A4. Measure the absorbance (OD) value of each well at a wavelength of 490 nm and calculate the cell viability using the following formula:

[0131]

[0132] The experimental results are shown in Table 3;

[0133] Table 3 Cell viability data

[0134]

[0135] The cell survival rates in Examples 1-5 were all above 90%, indicating that the antiglycation and antioxidant compositions provided by the present invention have no toxic effects on cells and have high safety.

[0136] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or alterations to the above-disclosed technical content to create equivalent embodiments without departing from the scope of the present invention. Any simple modifications, equivalent changes and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the scope of the present invention.

Claims

1. An anti-glycation and antioxidant composition, characterized in that, The composition comprises the following components: compound plant fermentation products, lecithin, and vitamin E; The compound plant fermentation product is obtained by fermenting mulberry leaf extract, amla extract, golden top pine mushroom powder and prickly pear powder as compound plant fermentation raw materials and inoculating compound bacteria. The compound bacteria consist of *Helicobacter schrenckii* and *Pediococcus pentosaceus* in a volume ratio of 1:1-2. The preservation number of *Helicobacter schrenckii* is CGMCC NO.1.2513, and the preservation number of *Pediococcus pentosaceus* is CGMCC NO.1.7665.

2. The composition according to claim 1, characterized in that, The compound plant fermentation raw materials include the following raw materials in weight percentage: 8%-12% mulberry leaf extract, 10%-15% amla extract, 5%-8% golden top pine ear powder, and 6%-10% prickly pear powder.

3. The composition according to claim 1 or 2, characterized in that, The compound plant fermentation raw material is prepared from the following raw materials in weight percentage: 10% mulberry leaf extract, 13% amla extract, 7% pine nut powder, 8% prickly pear powder and the remainder deionized water.

4. The composition according to claim 1, characterized in that, The amount of the compound bacteria inoculated is 3-5% of the weight of the compound plant fermentation raw materials.

5. The composition according to claim 1, characterized in that, The compound bacteria consist of Helenweissella and Pediococcus pentosaceus in a volume ratio of 1:1.

3.

6. The composition according to claim 1, characterized in that, The viable count of the *Helenweissella* strain was 4 × 10⁻⁶. 8 CFU / mL, the viable count of the *Pediococcus pentosaceus* was 4 × 10⁻⁶. 8 CFU / mL.

7. A method for preparing the composition according to any one of claims 1-6, characterized in that, Specifically, the following steps are included: A1. Weigh each component of the compound plant fermentation raw material according to the formula, add deionized water and mix evenly to obtain the fermentation substrate; A2. Inoculate the activated compound bacterial solution into the fermentation substrate for fermentation. After fermentation, filter and concentrate the solution, then add lecithin and vitamin E, and homogenize to obtain the fermentation composition.

8. The method for preparing the composition according to claim 7, characterized in that, In step A2, the fermentation temperature is 30-35℃ and the fermentation time is 24-48h.

9. The use of the composition according to any one of claims 1-6 or the composition prepared by the preparation method according to any one of claims 7-8 in the preparation of anti-glycation and antioxidant products.

10. The application as described in claim 9, characterized in that, The dosage forms of the product include tablets, powders, pills, liquids, or capsules.

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