A composition, microcapsule powder and preparation method, application and product of fat-burning weight loss efficacy

Abstract

The application discloses a composition with fat-burning and weight-reducing effects, microcapsule powder and a preparation method, application and product thereof, and belongs to the technical field of health products.The composition comprises fat-burning composition 1 and fat-burning composition 2;the fat-burning composition 1 comprises 6-9 parts of tea tree pollen and 4-6 parts of concentrated mango powder by weight;the fat-burning composition 2 comprises 2-4 parts of black garlic powder and 3-5 parts of black ginger powder by weight;the fat-burning composition 1 is prepared by ultrasonic extraction of tea tree pollen and concentrated mango powder with an ethanol solution;the fat-burning composition 2 is prepared by fermentation of black garlic powder and black ginger powder with compound bacteria;the compound bacteria are plant lactobacillus and saccharomyces cerevisiae with a mass ratio of 1:1-1.5.The composition is prepared by selecting tea tree pollen, concentrated mango powder, black garlic powder and black ginger powder as ingredients, improving the preparation process, significantly improving the utilization rate of active ingredients, and realizing the significant enhancement of the lipid-lowering effect by means of the synergistic effect of multiple components and multiple targets.

Description

Technical Field

[0001] This invention belongs to the field of health product technology, specifically relating to a composition, microcapsule powder, preparation method, application, and product with fat-burning and weight-loss effects. Background Technology

[0002] With changes in modern lifestyles, hyperlipidemia and related metabolic diseases have become a global public health problem. Non-alcoholic fatty liver disease (NAFLD), as the hepatic manifestation of metabolic syndrome, has an increasing incidence rate and has become one of the main causes of chronic liver disease. Excessive lipid accumulation in hepatocytes is a core pathological feature in the pathogenesis of NAFLD; therefore, finding natural active substances that can effectively inhibit lipid accumulation in hepatocytes has significant scientific and clinical value.

[0003] Chinese patent CN110801026A (published on February 18, 2020) discloses a dietary fiber complex nutrient with weight loss and lipid-lowering effects, which includes tea tree pollen, oat flour, psyllium husk powder, chia seed oil, cranberry concentrate, sodium citrate, annatto, tricalcium phosphate, konjac flour, dandelion extract, corn silk extract, resistant dextrin, L-arabinose, and mulberry leaf extract, indicating that tea tree pollen has lipid-lowering effects.

[0004] Chinese patent CN121312821A (published on January 13, 2026) discloses a composition comprising ginger and green tea microcapsule powder, raspberry ketone, mango concentrate powder, citrus fruit powder, and seaweed powder. This composition can directly act on key enzymes in fat degradation, enhancing the fat-reducing effect. This indicates that mango concentrate powder has fat-reducing efficacy.

[0005] Chinese patent CN119655322A (publication date March 21, 2025) discloses an instant sugar-controlling and fat-reducing solid milk tea, comprising tea powder, milk powder, freeze-dried ginger powder, coconut oil powder, flaxseed diglyceride oil powder, stevia powder and freeze-dried fruit powder, indicating that ginger has the effect of reducing lipids.

[0006] Black garlic is a food product made from garlic through high-temperature and high-humidity fermentation. It contains active chemical components such as melanoidins, proteins, polyphenols, sulfur-containing compounds, and polysaccharides, and has high nutritional value, which has gradually gained popularity in recent years. Studies have shown that black garlic has pharmacological effects such as anti-oxidation, anti-inflammation, anti-tumor, and lipid-lowering effects (Journal article: Guan Mengyao et al., Research progress on the medicinal and edible functions of black garlic [J], Food and Drug, 2024, Vol. 26, No. 1, pp. VIII-XIV).

[0007] Currently, research on the lipid-lowering activity of black garlic powder combined with tea tree pollen, concentrated mango powder, and black ginger is lacking. The applicant's research found that simply grinding and mixing these raw materials results in insufficient release and low utilization of active ingredients, leading to poor lipid-lowering effects and failing to meet market demand for highly effective, natural fat-burning and lipid-lowering health products. Summary of the Invention

[0008] To address the aforementioned technical problems, this invention provides a composition with fat-burning and weight-loss effects, its preparation method, and its application. The composition is formulated with tea tree pollen, concentrated mango powder, black garlic powder, and black ginger powder. Through improvements in the preparation process, the utilization rate of active ingredients is significantly increased. By leveraging the synergistic effect of multiple components and multiple targets, the lipid-lowering effect is greatly enhanced.

[0009] Terminology Explanation: Unless otherwise defined, all technical terms used herein have the same meanings as commonly understood by one of ordinary skill in the art. Unless otherwise stated, all patents, patent inventions, and publications cited in this document are incorporated herein by reference in their entirety. If multiple definitions exist for terms, the definition provided in this chapter shall prevail.

[0010] It should be understood that the above brief description and the following detailed description are exemplary and for illustrative purposes only, and do not limit the subject matter of the invention in any way. In this invention, the singular is used in conjunction with the plural unless otherwise specifically stated. It should also be noted that, unless otherwise stated, the use of “or” or “or” means “and / or”. Furthermore, the use of the term “comprising” and other forms such as “including,” “containing,” and “contains” are not limiting.

[0011] Definitions of standard chemical terms can be found in reference textbooks or reference books.

[0012] Unless specifically defined herein, the use of all commercially available products herein employs standard techniques. For example, it may be carried out using the manufacturer's instructions for use with the kit, or in accordance with methods known in the art or the description of this invention. The techniques and methods described herein can generally be implemented according to conventional methods well known in the art, based on the descriptions in the various summary and more specific documents cited and discussed in this specification.

[0013] As used herein, the term "room temperature" refers to ambient temperature, ranging from about 10°C to about 40°C. In some embodiments, "room temperature" refers to a temperature ranging from about 20°C to about 30°C; in other embodiments, "room temperature" refers to a temperature ranging from about 25°C to about 30°C; and in still other embodiments, "room temperature" refers to 10°C, 15°C, 20°C, 25°C, 30°C, 35°C, 40°C, etc.

[0014] The term "pulverization" as used in this article refers to the process of breaking down solid lumps or granules into fine particles, fine powders, or ultrafine powders through physical or mechanical methods such as impact, shearing, grinding, and shock. This process can disrupt the material's structure, increase its specific surface area, and improve its dissolution and extraction efficiency. Among these, mechanical pulverization, air jet milling, ultrafine pulverization, cryogenic pulverization, or grinding pulverization are all conventional pulverization methods in this field.

[0015] As used in this article, "solid-liquid separation" refers to the process of separating a mixture containing solid particles and liquid components into independent solid and liquid phases using physical or mechanical methods. This process can remove solid impurities, cell debris, precipitates, microbial aggregates, and other solid substances from the mixture to obtain a clear liquid phase or collect the target solid product. It is a commonly used separation and purification method in cell culture, sample pretreatment, and reagent preparation. Centrifugation, filtration, sedimentation, and pressure filtration are all conventional solid-liquid separation methods in this field.

[0016] As used in this article, "concentration" refers to the process of removing water and low-boiling-point volatile solvents from a solution, extract, or material system using physical separation methods such as heating evaporation, reduced pressure evaporation, membrane separation, or extraction, thereby increasing the content of effective components, reducing the system volume, and improving the material concentration. Among these, atmospheric pressure concentration, reduced pressure concentration, thin-film concentration, rotary concentration, or membrane concentration are all conventional concentration methods in this field.

[0017] As used in this article, "drying" refers to the process of removing free moisture, solvents, and volatile components from the interior and surface of materials, raw materials, pharmaceutical intermediates, or equipment through physical or thermal methods such as volatilization and evaporation, thereby reducing the moisture content of the material, inhibiting microbial growth, and improving the stability and shelf life of the material. Atmospheric pressure drying, reduced pressure drying, vacuum drying, freeze drying, and hot air drying are all conventional drying methods in this field.

[0018] The term "inoculation" as used in this article refers to the core operation of transferring a target cell line, cell suspension, or cell-containing biological material into a suitable cell culture vessel and cell culture medium under strictly aseptic conditions, providing a suitable environment for cell growth, proliferation, differentiation, or expression, and initiating the in vitro cell culture process. Static plate inoculation, shake-flask suspension inoculation, and bioreactor inoculation are all routine cell inoculation methods in this field.

[0019] As used in this article, "sterilization" refers to the process of killing or removing all forms of microorganisms, including bacteria, fungi, viruses, mycoplasma, chlamydia, and highly resistant bacterial spores and fungal spores, from the environment, instruments, and reagents using physical or chemical methods, so as to bring the object to a sterile state. High-pressure steam sterilization, dry heat sterilization, filtration sterilization, or radiation sterilization are all conventional sterilization methods in this field.

[0020] The term "microencapsulated powder" as used in this article refers to a powdered solid preparation obtained by encapsulating a functional core material with natural or polymeric wall materials through processes such as encapsulation, emulsification, spray drying, and coagulation, forming tiny capsule-like particles, followed by drying and sieving. Microencapsulated powder can achieve sustained-release and controlled-release of the core material, isolation of odors, isolation from light and oxygen, improved stability and water solubility, reduced irritation, and extended shelf life. Spray-dried microencapsulated powder, coagulated microencapsulated powder, fluidized bed microencapsulated powder, emulsified gel microencapsulated powder, or nano-microencapsulated powder are all conventional microencapsulated powder categories in this field.

[0021] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides: a composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2; The fat-burning composition 1, by weight, comprises 6-9 parts tea tree pollen and 4-6 parts concentrated mango powder; The fat-burning composition 2, by weight, includes 2-4 parts of black garlic powder and 3-5 parts of black ginger powder.

[0022] In some embodiments, the amount of tea pollen used, by weight, is 6 parts, 7 parts, 8 parts, 9 parts, or a range derived thereof.

[0023] In some embodiments, the concentrated mango powder is used in the amount of 4 parts, 5 parts, 6 parts, or a range derived thereof, by weight.

[0024] In some implementations, the amount of black garlic powder used is 2 parts, 3 parts, 4 parts, or a range derived thereof, by weight.

[0025] In some embodiments, the amount of black ginger powder used is 3 parts, 4 parts, 5 parts, or a range derived thereof, by weight.

[0026] In some embodiments, the fat-burning composition 1 is prepared by ultrasonic extraction of tea tree pollen and concentrated mango powder using an ethanol solution.

[0027] In some embodiments, the fat-burning composition 2 is prepared by fermentation of black garlic powder and black ginger powder with compound bacteria.

[0028] In some embodiments, the mass ratio of the fat-burning composition 1 to the fat-burning composition 2 is 3:1-3, for example 3:1, 3:2, 3:3, or a range derived therefrom.

[0029] Secondly, the present invention also provides a method for preparing the composition, comprising the following steps: S1. The prescribed amounts of tea tree pollen and concentrated mango powder are pulverized and mixed evenly to obtain a mixed powder; S2. The mixed powder was extracted with ethanol solution by ultrasonication, followed by solid-liquid separation. The clear liquid was concentrated and dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula, mix them evenly, add water to obtain a mixture; S4. The mixed liquid is fermented, and the fermentation liquid is concentrated, sterilized, separated from the solid liquid, and dried to obtain fat-burning composition 2.

[0030] In some embodiments, the mass fraction of the ethanol solution in step S2 is 5%-50%, for example, selectable as 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, or ranges derived therefrom.

[0031] In some implementations, the ultrasonic extraction in step S2 is: Power ranges from 800 to 1500W, and time ranges from 30 to 90 minutes; for example, power options include 800W, 900W, 1000W, 1100W, 1200W, 1300W, 1400W, 1500W, or a range derived therefrom; time options include 30 minutes, 40 minutes, 50 minutes, 60 minutes, 70 minutes, 80 minutes, 90 minutes, or a range derived therefrom. The ultrasonic extraction is performed at least once, preferably 1-4 times, and more preferably 2-3 times. The amount of ethanol solution added each time is 4 to 10 times the weight of the mixed powder, for example, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, or a range derived therefrom.

[0032] In some embodiments, the concentration in step S2 is to concentrate at 40-60°C until the weight ratio of the medicinal material to the concentrate is 1:0.7-0.9; for example, it can be selected as 1:0.7, 1:0.8, 1:0.9, or a range derived therefrom.

[0033] In some implementations, the ratio of black garlic powder and black ginger powder to water in step S3 is 1:15-50 (g / mL); for example, it can be selected as 1:15, 1:16, 1:17, 1:18, 1:19, 1:20, 1:21, 1:22, 1:23, 1:24, 1:25, 1:26, 1:27, 1:28, 1:29, 1:30, 1:31, 1:32, 1:33, 1:34, 1:35, 1:36, 1:37, 1:38, 1:39, 1:40, 1:41, 1:42, 1:43, 1:44, 1:45, 1:46, 1:47, 1:48, 1:49, 1:50, or a range derived therefrom.

[0034] In some implementations, the fermentation described in step S4 is as follows: inoculating the mixture with a compound of bacteria, fermenting at pH 4.5-5.5 and 25-35°C for 42-54 hours.

[0035] Preferably, the compound bacteria are *Lactobacillus plantarum* and *Saccharomyces cerevisiae*. More preferably, the mass ratio of *Lactobacillus plantarum* to *Saccharomyces cerevisiae* is 1:1-1.5, for example, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, or a range derived therefrom.

[0036] Preferably, the inoculation amount of the compound bacteria is 1%-5% of the mass of the mixture, for example, 1%, 2%, 3%, 4%, 5%, or a range derived therefrom.

[0037] In some embodiments, the concentration in step S4 is to concentrate to 75%-85% of the original fermentation broth mass, for example, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, or a range derived therefrom.

[0038] The term "pulverization" in this invention refers to pulverizing through a 100-200 mesh sieve, preferably through a 200 mesh sieve.

[0039] The "solid-liquid separation" involved in this invention refers to centrifugation or filtration. For example, centrifugation is centrifugation at 3000-5000 rpm for 5-15 min; the filtration membrane has a pore size of 0.45 μm.

[0040] The "sterilization" involved in this invention refers to sterilization at 121°C for 15-25 minutes, preferably 20 minutes.

[0041] Thirdly, the present invention also provides: a microcapsule powder with fat-burning and weight-loss effects, comprising the fat-burning composition 1, the fat-burning composition 2, diglycerides and seaweed powder.

[0042] In some embodiments, the mass ratio of the fat-burning composition 1, the fat-burning composition 2, the diglyceride, and the seaweed powder is 3:1-3:0.8-1.2:8-12, for example, 3:1:0.8:8, 3:1:0.8:9, 3:1:0.8:10, 3:1:0.8:11, 3:1:0.8:12, 3:1:0.9:8, 3:1:0.9:9, 3:1:0.9:10, 3:1:0.9:11, 3:1:0.9:12, or 3:1:1. 0:8, 3:1:1.0:9, 3:1:1.0:10, 3:1:1.0:11, 3:1:1.0:12, 3:1:1.1:8, 3:1:1.1:9, 3:1:1.1:10, 3:1:1.1:11, 3:1:1.1:12, 3:2:0.8:8, 3:2:0.8:9, 3:2:0.8:10, 3:2:0.8:11, 3:2:0.8:12, 3:2:0.9:8, 3:2:0.9:9, 3:2: 0.9:10, 3:2:0.9:11, 3:2:0.9:12, 3:2:1.0:8, 3:2:1.0:9, 3:2:1.0:10, 3:2:1.0:11, 3:2:1.0:12, 3:2:1.1:8, 3:2:1.1:9, 3:2:1.1:10, 3:2:1.1:11, 3:2:1.1:12, 3:3:0.8:8, 3:3:0.8:9, 3:3:0.8:10, 3:3:0.8:11 3:3:0.8:12, 3:3:0.9:8, 3:3:0.9:9, 3:3:0.9:10, 3:3:0.9:11, 3:3:0.9:12, 3:3:1.0:8, 3:3:1.0:9, 3:3:1.0:10, 3:3:1.0:11, 3:3:1.0:12, 3:3:1.1:8, 3:3:1.1:9, 3:3:1.1:10, 3:3:1.1:11, 3:3:1.1:12, or ranges derived therefrom.

[0043] Fourthly, the present invention also provides a method for preparing the microcapsule powder, comprising the following steps: mixing the fat-burning composition 1, the fat-burning composition 2, diglycerides and seaweed powder.

[0044] Fifthly, the present invention provides the application of the composition, the composition prepared by the preparation method, the microcapsule powder or the microcapsule powder prepared by the preparation method in the preparation of fat-burning products.

[0045] The product described in this invention is a health supplement.

[0046] In a sixth aspect, the present invention provides a fat-burning product made from the composition, the composition prepared by the preparation method, the microcapsule powder or the microcapsule powder prepared by the preparation method, and excipients acceptable in health products.

[0047] The present invention has at least the following beneficial effects: This invention selects tea tree pollen, concentrated mango powder, black garlic powder, and black ginger powder for formulation. The black garlic and black ginger powders are fermented using a compound microbial fermentation process. On one hand, the enzymes produced by the microorganisms decompose plant cell walls, improving the dissolution of active ingredients. On the other hand, microbial metabolism generates various secondary active substances, enhancing the lipid-lowering activity of the raw materials themselves. The tea tree pollen and concentrated mango powder are enriched with active ingredients through ultrasonic extraction with ethanol. After combining these two types of process products, the utilization rate of active ingredients is significantly improved. Each component exerts a complementary and synergistic effect across multiple targets, significantly enhancing the composition's effect of inhibiting lipid accumulation and lowering blood lipids. Attached Figure Description

[0048] Figure 1 Image showing the results of staining lipid droplets with Oil Red O. Detailed Implementation

[0049] The following non-limiting embodiments are intended to enable those skilled in the art to gain a more comprehensive understanding of the present invention, but do not limit the invention in any way. The following content is merely an exemplary description of the scope of protection claimed by the present invention, and those skilled in the art can make various changes and modifications to the present invention based on the disclosed content, and such changes should also fall within the scope of protection claimed by the present invention.

[0050] All numerical values ​​or expressions relating to component amounts, process conditions, etc., used in this invention shall be understood to be modified by the word "about" in all cases. When referring to a quantity or range of values, the quantity or range is an approximation within experimental variability (or within statistical experimental error). In this invention, the term "about" shall have the meaning of being within 10%, preferably within 5%, of the specified value or range.

[0051] The present invention will be further described below by way of specific embodiments. Unless otherwise specified, all instruments, devices, equipment, reagents, products, etc., used in the embodiments of the present invention are obtained through conventional commercial means.

[0052] Unless otherwise specified, percentages in this invention refer to mass percentages, temperatures refer to room temperature, and solvents refer to water.

[0053] For example, the raw material information in this embodiment is as follows: Tea tree pollen, purchased from Jiangxi Sidarui Biotechnology Co., Ltd.; Product No.: 202602021P; Concentrated mango powder, purchased from Jiangxi Sidarui Biotechnology Co., Ltd.; Product No.: C00020; Black garlic powder, purchased from Jiangxi Sidarui Biotechnology Co., Ltd.; Product No.: C120606-03; Black ginger powder, purchased from Jiangxi Sidarui Biotechnology Co., Ltd.; Product No.: C00001; Diglycerides, purchased from Shandong Tianjiao Biotechnology Co., Ltd.; Product No.: DAG-YS60; Lactobacillus plantarum, purchased from Qingdao Novo Nordisk Biotechnology Co., Ltd.; Product No.: GBW1051; Brewing yeast, purchased from Angel Yeast Co., Ltd.; product number: 85001505.

[0054] HepG2 cells (human liver cancer cell line) were obtained from the China Center for Type Culture Collection (CCTCC). The passage number was controlled between 10 and 30 to avoid using cells with excessively high passage numbers. The cell morphology was epithelial-like adherent growth, polygonal, with a high nucleocytoplasmic ratio. The culture conditions were 37°C, 5% CO2, and saturated humidity.

[0055] The complete culture medium was DMEM high glucose medium containing 10 v / v% fetal bovine serum FBS, 1 v / v% penicillin-streptomycin antibiotics, PBS buffer, pH 7.4.

[0056] The preparation method of free fatty acid (FFA) inducer is as follows: (1) Weigh an appropriate amount of sodium oleate, dissolve it in anhydrous ethanol, and prepare a 100 mM stock solution; (2) Weigh an appropriate amount of sodium palmitate, dissolve it in anhydrous ethanol, and prepare a 100 mM stock solution; (3) Dilute the stock solution with complete culture medium to make the final fatty acid concentration the required concentration.

[0057] (4) Mix oleic acid and palmitic acid in a molar ratio of 1:1 to prepare a working solution with a total concentration of 1.0 mM FFA.

[0058] (5) Filtration and sterilization (0.22 μm filter membrane), prepare and use immediately.

[0059] Example 1 A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 1:1; The fat-burning composition 1 comprises, by weight, 9 parts tea tree pollen and 4 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 4 parts black garlic powder and 3 parts black ginger powder.

[0060] The preparation method of the composition includes the following steps: S1. The tea tree pollen and concentrated mango powder of the formula amount are pulverized through a 200-mesh sieve and mixed evenly to obtain a mixed powder; S2. The mixed powder was placed in an extraction tank and ultrasonically extracted twice (power 1200W), 1 hour each time. For the first extraction, 40% ethanol solution with a mass fraction of 8 times the weight of the mixed powder was added. For the second extraction, 10% ethanol solution with a mass fraction of 6 times the weight of the mixed powder was added. The extracts were combined and centrifuged at 4000 rpm for 10 minutes. The supernatant was concentrated to a weight ratio of 1:0.8 between the medicinal material and the concentrated liquid. The mixture was then spray-dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula into powder through a 200-mesh sieve, mix them evenly, and add water at a material-to-liquid ratio of 1:15 (g / mL) to obtain a mixture. S4. Mixed liquor fermentation: The inoculum of compound bacteria (Lactobacillus plantarum and Saccharomyces cerevisiae in a mass ratio of 1:1) is 5% of the mass of the mixed liquor. The fermentation temperature is 25℃, the fermentation pH is 4.5-5.5, the sterile air ventilation rate is 5L / min, the rotation speed is 200rpm, and the fermentation time is 54h. After fermentation, the fermentation broth is concentrated under reduced pressure (40℃, -0.06MPa) to 80% of the original fermentation broth mass, sterilized at 121℃ for 20min, filtered through a 0.45μm membrane, and the clear liquid is freeze-dried to obtain fat-burning composition 2.

[0061] S5. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0062] Example 2 A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 3:1; The fat-burning composition 1 comprises, by weight, 6 parts tea tree pollen and 6 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 2 parts black garlic powder and 5 parts black ginger powder.

[0063] The preparation method of the composition includes the following steps: S1. The tea tree pollen and concentrated mango powder of the formula amount are pulverized through a 200-mesh sieve and mixed evenly to obtain a mixed powder; S2. The mixed powder was placed in an extraction tank and ultrasonically extracted twice (power 1200W), 1 hour each time. For the first extraction, 50% ethanol solution with a mass fraction of 10 times the weight of the mixed powder was added. For the second extraction, 5% ethanol solution with a mass fraction of 4 times the weight of the mixed powder was added. The extracts were combined and centrifuged at 4000 rpm for 10 minutes. The supernatant was concentrated to a weight ratio of 1:0.8 between the medicinal material and the concentrated liquid. The mixture was then spray-dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula into powder through a 200-mesh sieve, mix them evenly, and add water at a material-to-liquid ratio of 1:50 (g / mL) to obtain a mixture. S4. Fermentation of the mixed liquid: The inoculum of compound bacteria (Lactobacillus plantarum and Saccharomyces cerevisiae in a mass ratio of 1:1.5) is 1% of the mass of the mixed liquid. The fermentation temperature is 35℃, the fermentation pH is 4.5-5.5, the sterile air ventilation rate is 5L / min, the rotation speed is 200rpm, and the fermentation time is 42h. After fermentation, the fermentation liquid is concentrated under reduced pressure (40℃, -0.06MPa) to 80% of the original fermentation liquid mass, sterilized at 121℃ for 20min, filtered through a 0.45μm membrane, and the clear liquid is freeze-dried to obtain fat-burning composition 2.

[0064] S5. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0065] Example 3 A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 3:2; The fat-burning composition 1 comprises, by weight, 8 parts tea tree pollen and 5 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 3 parts black garlic powder and 4 parts black ginger powder.

[0066] The preparation method of the composition includes the following steps: S1. The tea tree pollen and concentrated mango powder of the formula amount are pulverized through a 200-mesh sieve and mixed evenly to obtain a mixed powder; S2. The mixed powder was placed in an extraction tank and ultrasonically extracted twice (power 1200W), 1 hour each time. For the first extraction, 40% ethanol solution with a mass fraction of 10 times the weight of the mixed powder was added. For the second extraction, 10% ethanol solution with a mass fraction of 4 times the weight of the mixed powder was added. The extracts were combined and centrifuged at 4000 rpm for 10 minutes. The supernatant was concentrated to a weight ratio of 1:0.8 between the medicinal material and the concentrated liquid. The mixture was then spray-dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula into powder through a 200-mesh sieve, mix them evenly, and add water at a material-to-liquid ratio of 1:50 (g / mL) to obtain a mixture. S4. Mixed liquor fermentation: The inoculum of compound bacteria (Lactobacillus plantarum and Saccharomyces cerevisiae in a mass ratio of 1:1.2) is 1% of the mixed liquor mass. The fermentation temperature is 35℃, the fermentation pH is 4.5-5.5, the sterile air ventilation rate is 5L / min, the rotation speed is 200rpm, and the fermentation time is 50h. After fermentation, the fermentation broth is concentrated under reduced pressure (40℃, -0.06MPa) to 80% of the original fermentation broth mass, sterilized at 121℃ for 20min, filtered through a 0.45μm membrane, and the clear liquid is freeze-dried to obtain fat-burning composition 2.

[0067] S5. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0068] Comparative Example 1 The only difference between this comparative example and Example 3 is that the preparation method of the composition is different.

[0069] Specifically: A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 3:2; The fat-burning composition 1 comprises, by weight, 8 parts tea tree pollen and 5 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 3 parts black garlic powder and 4 parts black ginger powder.

[0070] The preparation method of the composition includes the following steps: S1. The tea tree pollen and concentrated mango powder of the specified formula are pulverized through a 200-mesh sieve and mixed evenly to obtain fat-burning composition 1; S2. The specified amounts of black garlic powder and black ginger powder are pulverized through a 200-mesh sieve and mixed evenly to obtain fat-burning composition 2.

[0071] S3. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0072] Comparative Example 2 The only difference between this comparative example and Example 3 is that the compound bacteria used in the preparation method of the composition is *Lactobacillus plantarum*.

[0073] Specifically: A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 3:2; The fat-burning composition 1 comprises, by weight, 8 parts tea tree pollen and 5 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 3 parts black garlic powder and 4 parts black ginger powder.

[0074] The preparation method of the composition includes the following steps: S1. The tea tree pollen and concentrated mango powder of the formula amount are pulverized through a 200-mesh sieve and mixed evenly to obtain a mixed powder; S2. The mixed powder was placed in an extraction tank and ultrasonically extracted twice (power 1200W), 1 hour each time. For the first extraction, 40% ethanol solution with a mass fraction of 10 times the weight of the mixed powder was added. For the second extraction, 10% ethanol solution with a mass fraction of 4 times the weight of the mixed powder was added. The extracts were combined and centrifuged at 4000 rpm for 10 minutes. The supernatant was concentrated to a weight ratio of 1:0.8 between the medicinal material and the concentrated liquid. The mixture was then spray-dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula into powder through a 200-mesh sieve, mix them evenly, and add water at a material-to-liquid ratio of 1:50 (g / mL) to obtain a mixture. S4. Fermentation of the mixed liquid: The inoculum amount of *Lactobacillus plantarum* was 1% of the mass of the mixed liquid. The fermentation temperature was 35℃, the fermentation pH was 4.5-5.5, the sterile air ventilation rate was 5L / min, the rotation speed was 200rpm, and the fermentation time was 50h. After fermentation, the fermentation liquid was concentrated under reduced pressure (40℃, -0.06MPa) to 80% of the original fermentation liquid mass. It was sterilized at 121℃ for 20min, filtered through a 0.45μm membrane, and the clear liquid was freeze-dried to obtain fat-burning composition 2.

[0075] S5. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0076] Comparative Example 3 The only difference between this comparative example and Example 3 is that: black garlic powder and black ginger powder were prepared by ultrasonic extraction with ethanol solution; and tea tree pollen and concentrated mango were prepared by fermentation with compound bacteria.

[0077] Specifically: A composition with fat-burning and weight-loss effects, comprising fat-burning composition 1 and fat-burning composition 2 in a mass ratio of 3:2; The fat-burning composition 1 comprises, by weight, 8 parts tea tree pollen and 5 parts concentrated mango powder; The fat-burning composition 2 comprises, by weight, 3 parts black garlic powder and 4 parts black ginger powder.

[0078] The preparation method of the composition includes the following steps: S1. Grind the black garlic powder and black ginger according to the formula into powder, pass them through a 200-mesh sieve, and mix them evenly to obtain a mixed powder; S2. The mixed powder was placed in an extraction tank and ultrasonically extracted twice (power 1200W), 1 hour each time. For the first extraction, 40% ethanol solution with a mass fraction of 10 times the weight of the mixed powder was added. For the second extraction, 10% ethanol solution with a mass fraction of 4 times the weight of the mixed powder was added. The extracts were combined and centrifuged at 4000 rpm for 10 minutes. The supernatant was concentrated to a weight ratio of 1:0.8 between the medicinal material and the concentrated liquid. The mixture was then spray-dried to obtain fat-burning composition 1. S3. The tea tree pollen and concentrated mango in the formula amount are crushed through a 200-mesh sieve, mixed evenly, and water is added at a material-to-liquid ratio of 1:50 (g / mL) to obtain a mixed solution. S4. Mixed liquor fermentation: The inoculum of compound bacteria (Lactobacillus plantarum and Saccharomyces cerevisiae in a mass ratio of 1:1.2) is 1% of the mixed liquor mass. The fermentation temperature is 35℃, the fermentation pH is 4.5-5.5, the sterile air ventilation rate is 5L / min, the rotation speed is 200rpm, and the fermentation time is 50h. After fermentation, the fermentation broth is concentrated under reduced pressure (40℃, -0.06MPa) to 80% of the original fermentation broth mass, sterilized at 121℃ for 20min, filtered through a 0.45μm membrane, and the clear liquid is freeze-dried to obtain fat-burning composition 2.

[0079] S5. Simply mix fat-burning composition 1 and fat-burning composition 2 in the specified proportions.

[0080] Example 4 A microcapsule powder with fat-burning and weight-loss effects The microcapsule powder consists of fat-burning composition 1, fat-burning composition 2, diglycerides, and seaweed powder in a mass ratio of 3:1:0.8:12.

[0081] The preparation method is as follows: Mix fat-burning composition 1, fat-burning composition 2, diglycerides, and seaweed powder evenly according to the specified proportions. The fat-burning composition 1 and fat-burning composition 2 were prepared in Example 3.

[0082] Example 5 A microcapsule powder with fat-burning and weight-loss effects The microcapsule powder consists of fat-burning composition 1, fat-burning composition 2, diglycerides, and seaweed powder in a mass ratio of 3:3:1.2:8.

[0083] The preparation method is as follows: Mix fat-burning composition 1, fat-burning composition 2, diglycerides and seaweed powder evenly according to the proportion; The fat-burning composition 1 and fat-burning composition 2 were prepared in Example 3.

[0084] Example 6 A microcapsule powder with fat-burning and weight-loss effects The microcapsule powder consists of fat-burning composition 1, fat-burning composition 2, diglycerides, and seaweed powder in a mass ratio of 3:2:1:10.

[0085] The preparation method is as follows: Mix fat-burning composition 1, fat-burning composition 2, diglycerides and seaweed powder evenly according to the proportion; The fat-burning composition 1 and fat-burning composition 2 were prepared in Example 3.

[0086] Example 1: Safety Evaluation The cytotoxicity assay (CCK-8 assay) is a widely used method for detecting cell viability. WST-8 is reduced by dehydrogenases in the mitochondria of live cells in the presence of an electron-coupling reagent, producing an orange-yellow formazan dye. The amount of formazan produced is directly proportional to the number of live cells; cell viability can be calculated by measuring the absorbance.

[0087] 1. Cell inoculation (1) Preparation of cell suspension: Collect HepG2 cells in the logarithmic growth phase, count them, and adjust the cell density to 5×10⁻⁶. 4 cells / mL; (2) Cell seeding: Add 100 μL of cell suspension (approximately 5 × 10⁶ cells / well) to each well of a 96-well plate.3 cells / pores); (3) Blank control: Set up wells with only complete culture medium and no cells as blank control (set up 6 replicates). (4) Culture: Place the 96-well plate in an incubator and culture at 37°C and 5% CO2 for 24 hours to allow the cells to adhere to the plate.

[0088] 2. Treatment of the test substance (1) Preparation of test substance: Prepare a series of concentrations of test substance with complete culture medium, with concentration gradients of 0, 12.5, 25, 50, 100, 200, 400 and 800 μg / mL, and set up 6 replicates for each concentration; at the same time, set up a solvent control (containing cells, an equal volume of complete culture medium, and no test substance).

[0089] (2) Adding samples: Discard the old culture medium and add 100 μL of complete culture medium containing the corresponding concentration of the test substance to each well. Be careful to handle it gently and avoid disturbing the cells.

[0090] (3) Culture: Put the 96-well plate back into the incubator and continue to culture for 24 hours.

[0091] 3. CCK-8 detection (1) Prepare CCK-8 working solution: Mix CCK-8 reagent with complete culture medium in proportion according to the kit instructions.

[0092] (2) Add CCK-8: Discard the old culture medium and add 110 μL of CCK-8 working solution to each well.

[0093] (3) Incubation: Place the culture plate back into the incubator at 37°C and 5% CO2 for 30 minutes.

[0094] (4) Measure absorbance: Use an enzyme-linked immunosorbent assay (ELISA) reader to measure the absorbance (OD) of each well at 450 nm. Gently shake the culture plate before measurement to ensure uniform dye distribution.

[0095] 4. Data Processing (1) Calculate the corrected absorbance for each well: Corrected OD = Average OD of measured wells - Average OD of blank control wells.

[0096] (2) Calculate cell viability: Cell viability (%) = (OD corrected for test sample wells / OD corrected for solvent control wells) × 100%.

[0097] (3) Determine the safe concentration range: A concentration with a cell viability rate >90% is considered a non-toxic concentration.

[0098] 5. Results The compositions prepared in Examples 1-3 and Comparative Examples 1-3 all showed good safety in the range of 12.5 ug / mL to 200 ug / mL. The following description uses the cytotoxicity test results of the composition prepared in Example 3 as an example to illustrate the safety of the composition.

[0099] The cytotoxicity test results of the composition prepared in Example 3 are shown in Table 1.

[0100] Table 1

[0101] The results showed that the composition prepared in this invention has good safety in the range of 12.5 ug / mL to 200 ug / mL and can be used for subsequent experiments.

[0102] Example 2: Evaluation of lipid-lowering activity Free fatty acids (FFA) are a significant factor in causing hepatocyte steatosis. Exogenous FFA enters hepatocytes and is esterified into triglycerides by related enzymes. When FFA supply is excessive or triglyceride (TG) metabolism is impaired, TG accumulates in the cytoplasm as lipid droplets, leading to a steatosis phenotype. Oleic acid and palmitic acid are the two most abundant free fatty acids in the human body, and their mixture can effectively induce a HepG2 cell steatosis model.

[0103] 1. Cell inoculation (1) Preparation of cell suspension: Collect HepG2 cells in the logarithmic growth phase, count them, and adjust the cell density to 5×10⁻⁶. 4 cells / mL; (2) Cell seeding: Add 100 μL of cell suspension (approximately 5 × 10⁶ cells / well) to each well of a 96-well plate. 3 cells / pores); (3) Cultivation: Place in an incubator and incubate at 37°C and 5% CO2 for 24 hours. Discard the old culture medium.

[0104] 2. Model establishment and test substance intervention (1) Grouping Test group: each well was filled with complete culture medium containing FFA (0.5 mM oleic acid + 0.5 mM palmitic acid) and 0.05 mg / mL of the test substance (the composition prepared in the examples or comparative examples); Normal control group: Add complete culture medium to each well; Model group: Each well was filled with complete culture medium containing FFA (0.5 mM oleic acid + 0.5 mM palmitic acid); Positive control group: Each well was filled with complete culture medium containing FFA (0.5 mM oleic acid + 0.5 mM palmitic acid) and 10 µM simvastatin.

[0105] Each group has 3 duplicate holes.

[0106] (2) Cultivation: Place the culture plate in an incubator and incubate at 37°C and 5% CO2 for 24 hours.

[0107] 3. Observation using Oil Red O staining Cells were stained with Oil Red O after treatment. The steps are as follows: (1) Cell fixation: Discard the old culture medium, gently wash the cells twice with PBS, add 4v / v% paraformaldehyde fixative to each well, fix at room temperature for 30 minutes, or fix at 4°C overnight; (2) Washing: Discard the fixative and wash with PBS 3 times, 5 minutes each time; (3) Staining: Discard the PBS, add 0.3% Oil Red O working solution to each well, and stain at room temperature in the dark for 30 minutes. You can gently shake the well during the staining process to make the staining uniform.

[0108] (4) Differentiation: Discard the staining solution and use 60v / v% isopropanol to differentiate 2-3 times, each time for 30 seconds to 1 minute, until the background is clear and the lipid droplets are clearly visible.

[0109] (5) Counterstaining: Wash twice with PBS, add hematoxylin staining solution to counterstain cell nuclei for 1-2 minutes, and wash with PBS.

[0110] (6) Observation and photography: Under an inverted microscope, lipid droplets appear red and cell nuclei appear blue. Take typical field-of-view photos.

[0111] 4. Quantitative analysis of Oil Red O The quantitative analysis of Oil Red O follows these steps: (1) After staining, discard the PBS; (2) Add ethanol to each well; (3) Shake at room temperature for 10-15 minutes to completely wash away the dye; (4) Transfer the eluent to a 96-well plate; (5) Measure the absorbance at 510 nm using an enzyme-linked immunosorbent assay (ELISA) reader.

[0112] 5. Results Qualitative analysis of oil red O staining of lipid droplets, results are as follows: Figure 1 As shown.

[0113] As shown in the figure, under the microscope, compared with the normal control group, the model group cells were filled with a large number of dense, bright red lipid droplets, indicating that the intracellular fatty degeneration model was successfully constructed.

[0114] After drug administration intervention, compared with the model group, the area and staining depth of red lipid droplets in the cells of the positive control group and the Example 1-Example 3 groups showed a visible reduction, while the area and staining depth of red lipid droplets in the cells of the Comparative Example 1-Comparative Example 3 groups showed no significant change.

[0115] At the morphological level, the results directly confirm that the compositions prepared in Examples 1-3 have the ability to inhibit or reverse abnormal triglyceride deposition, while the compositions prepared in Comparative Examples 1-3 do not have the above-mentioned effect.

[0116] To eliminate subjective visual errors, this invention utilizes ethanol to elute the bound oil red dye and measures the absorbance at 510 nm. The results are shown in Table 2.

[0117] Table 2

[0118] The results showed that the OD value of the model group was significantly higher than that of the normal control group (P<0.01), indicating that the relative lipid content of cells increased significantly after modeling. After drug intervention, compared with the model group, the OD values ​​of the positive control group and the Example 1-Example 3 groups were significantly reduced (P<0.01 or 0.05), while there was no statistically significant difference in the OD values ​​of the Comparative Example 1-Comparative Example 3 groups (P>0.05). This indicates that the compositions prepared in Example 1-Example 3 can significantly reduce the relative lipid content in cells and have high lipid-lowering activity, while the compositions prepared in Comparative Example 1-Comparative Example 3 did not have the above effects.

[0119] Example 3: Determination of Triglyceride Content Triglycerides (TG) are hydrolyzed by lipoprotein lipase to produce glycerol and fatty acids. Glycerol is converted into glycerol-3-phosphate by glycerol kinase, which is then oxidized by glycerol phosphate oxidase to produce hydrogen peroxide (H2O2). H2O2 reacts with 4-aminoantipyrine and phenol under the action of peroxidase to produce red quinone compounds, which have a characteristic absorption peak at a wavelength of 500-550 nm. The absorbance is directly proportional to the TG content.

[0120] Following the steps outlined in Example 2, complete the cell seeding, model establishment, and test substance intervention. (1) Termination treatment: Discard the old culture medium and wash the cells twice with PBS pre-cooled to 4°C; (2) Cell collection: After adding PBS, scrape off the cells with a cell scraper, transfer the cell suspension to a centrifuge tube, centrifuge at 1000 rpm for 5 minutes, discard the supernatant, and add lysis buffer to resuspend the cell pellet. (3) Sample preparation: Sonicate on ice to completely lyse the cells. Take part of the lysate for BCA protein determination and the remaining lysate for TG determination. Triglyceride and BCA protein assays were performed according to the kit instructions.

[0121] Data processing: (1) Calculate TG Calculate the TG content (μg) of each sample based on the standard curve. TG (μg) = (OD of test group - OD of normal control group) ÷ slope of standard curve; (2) Calculate TG 绝对含量 TG 绝对含量 (μg / mg) = TG ÷ Protein content; (3) Calculate the inhibition rate TG inhibition rate (%) = [1 - (test group TG)] 绝对含量 - Normal control group TG 绝对含量 ) / (Model Group TG 绝对含量 - Normal control group TG 绝对含量 )]×100%.

[0122] The results are shown in Table 3.

[0123] Table 3

[0124] Triglycerides (TG) are a core quantitative biochemical indicator reflecting the pathological characteristics of non-alcoholic fatty liver disease. To further clarify the lipid-lowering efficacy of the composition of this invention, intracellular TG levels in each group were measured. 绝对含量 Experimental results showed that intracellular TG in the normal control group cells... 绝对含量 The value was 45.02±3.47 μg / mg. Under FFA induction, the intracellular TG in the model group cells was... 绝对含量 Significantly increased (P < 0.01); after drug intervention, intracellular TG in the positive control group and the groups of Examples 1-3 was significantly increased. 绝对含量 Significantly reduced (P < 0.01); intracellular TG in Comparative Example 1-Comparative Example 3 groups 绝对含量 The lack of statistical difference indicates that the compositions prepared in Examples 1-3 can penetrate the cellular lipid metabolism process, fundamentally and effectively intervening in and reducing the generation and accumulation of triglycerides; the compositions prepared in Comparative Examples 1-3 do not have the above-mentioned effects.

[0125] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, and is not intended to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions made by those skilled in the art to the technical solution of the present invention do not depart from the essence and scope of the technical solution of the present invention.

Claims

1. A composition with fat-burning and weight-loss effects, characterized in that, Includes fat-burning composition 1 and fat-burning composition 2; The fat-burning composition 1, by weight, comprises 6-9 parts tea tree pollen and 4-6 parts concentrated mango powder; The fat-burning composition 2, by weight, includes 2-4 parts of black garlic powder and 3-5 parts of black ginger powder; The fat-burning composition 1 is prepared by ultrasonic extraction of tea tree pollen and concentrated mango powder with ethanol solution; The fat-burning composition 2 is prepared by fermentation of black garlic powder and black ginger powder with compound bacteria. The compound bacteria consist of *Lactobacillus plantarum* and *Saccharomyces cerevisiae* in a mass ratio of 1:1-1.

5. The fermentation process is as follows: pH 4.5-5.5, fermentation at 25-35℃ for 42-54 hours; The mass ratio of fat-burning composition 1 to fat-burning composition 2 is 3:1-3.

2. The composition according to claim 1, characterized in that, The ethanol solution has a mass fraction of 5%-50%.

3. The composition according to any one of claims 1-2, characterized in that, The mass ratio of fat-burning composition 1 to fat-burning composition 2 is 1:

1.

4. A method for preparing the composition according to any one of claims 1-3, comprising the following steps: S1. The prescribed amounts of tea tree pollen and concentrated mango powder are pulverized and mixed evenly to obtain a mixed powder; S2. The mixed powder was extracted with ethanol solution by ultrasonication, followed by solid-liquid separation. The clear liquid was concentrated and dried to obtain fat-burning composition 1. S3. Grind the black garlic powder and black ginger powder according to the formula, mix them evenly, add water to obtain a mixture; S4. The mixed liquid is fermented, and the fermentation liquid is concentrated, sterilized, separated from the solid liquid, and dried to obtain fat-burning composition 2.

5. The preparation method according to claim 4, characterized in that, The ultrasonic extraction described in step S2 involves at least one ultrasonic extraction. Each time, the amount of ethanol solution added should be 4-10 times the weight of the mixed powder.

6. The preparation method according to claim 4, characterized in that, The ratio of black garlic powder and black ginger powder to water in step S3 is 1:15-50, g / mL; The inoculation amount of the compound bacteria is 1%-5% of the mass of the mixture.

7. A microcapsule powder with fat-burning and weight-loss effects, characterized in that, Includes the composition according to any one of claims 1-3 or the composition prepared by the preparation method according to any one of claims 4-6, diglycerides, and seaweed powder.

8. The microencapsulated powder according to claim 7, characterized in that, The mass ratio of the fat-burning composition 1, the fat-burning composition 2, diglycerides, and seaweed powder is 3:1-3:0.8-1.2:8-12.

9. A method for preparing the microcapsule powder according to any one of claims 7-8, characterized in that, The process includes the following steps: mixing the fat-burning composition 1, the fat-burning composition 2, diglycerides, and seaweed powder.

10. The use of the composition according to any one of claims 1-3 or the composition prepared by the preparation method according to any one of claims 4-6, the microcapsule powder according to any one of claims 7-8 or the microcapsule powder prepared by the preparation method according to claim 9 in the preparation of fat-burning products.

11. A fat-burning product, characterized in that, The product is made from the composition according to any one of claims 1-3 or the composition prepared by the preparation method according to any one of claims 4-6, the microcapsule powder according to any one of claims 7-8, the microcapsule powder prepared by the preparation method according to claim 9, and excipients acceptable in health products.

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