Lotus leaf black tea composite lipid-lowering substitute tea and preparation method thereof

Abstract

This invention discloses a lotus leaf and dark tea compound lipid-lowering substitute tea and its preparation method, belonging to the field of food processing technology. The method includes enzymatically hydrolyzed lotus leaf powder and highly active dark tea base material, which are then combined and subjected to directional fermentation under coupled conditions of ultrasound assistance and segmented temperature and humidity control. The substitute tea contains a nanoscale self-assembled inclusion structure spontaneously formed by non-covalent bonding of lotus leaf alkaloids, theabrownins, and tea polysaccharides. The enzymatically hydrolyzed lotus leaf powder is obtained by enzymatic hydrolysis of lotus leaf raw materials using a compound biological enzyme, and the highly active dark tea base material is obtained by co-fermentation and domestication of dark tea raw materials with specific probiotics. This invention achieves a significant enhancement of the intestinal-targeted sustained release of functional factors and synergistic lipid-lowering effects. Finally, by combining a low-temperature drying process, the microscopic morphology and high-activity component content are locked in, resulting in a functional substitute tea with both excellent lipid-lowering performance and superior sensory quality.

Description

Technical Field

[0001] This invention relates to the field of food processing technology, and in particular to a lotus leaf black tea compound lipid-lowering substitute tea and its preparation method. Background Technology

[0002] Lotus leaves and dark tea, as traditional medicinal and edible resources, have a broad application basis in regulating lipid metabolism. Lotus leaves are rich in lotus leaf alkaloids, flavonoids, and alkaloids, and are considered to have significant effects in inhibiting fat absorption and promoting lipid decomposition. Dark tea, known for its unique post-fermentation process, is rich in theaflavins, tea polysaccharides, and metabolites of *Aspergillus cristatus*, exhibiting good biological activity in improving intestinal flora structure and assisting in lipid reduction. Current technologies for the combined use of lotus leaves and dark tea mostly employ physical mixing and brewing, simple blending and pressing, or conventional co-fermentation processes. For example, some existing technologies directly mix dried lotus leaves (crushed) with raw dark tea leaves in a specific ratio for uniform pile fermentation or direct packaging into tea bags. These methods combine the flavors of the two raw materials to some extent, attempting to enhance the product's health benefits through a compounding effect. They also offer advantages in industrial production such as ease of operation and controllable costs, making them the mainstream preparation method in the current substitute tea market.

[0003] The dense cell wall structure of lotus leaves makes it difficult to completely break down the cell walls using conventional physical methods. This limits the dissolution rate of intracellular lotus leaf alkaloids and flavonoid active ingredients. Furthermore, the bioavailability of large-molecule polysaccharides from lotus leaves that have not undergone targeted enzymatic hydrolysis is low in the human body. During traditional co-fermentation, due to the lack of precise spatiotemporal control over the oxidative polymerization and molecular assembly stages, lotus leaf alkaloids are prone to degradation or non-specific binding and precipitation with tea polyphenols under strong oxidative conditions, making it difficult to form a stable molecular-level synergistic system with theaflavins and tea polysaccharides produced during the fermentation of dark tea. This lack of microscopic interaction results in the active ingredients in the final product often being distributed in a free state. This not only limits the full realization of the synergistic lipid-lowering effect of lotus leaf alkaloids and theaflavins, but also leaves a residual astringent taste characteristic of lotus leaves in the product's flavor, failing to achieve deep integration and stable maintenance of flavor substances and functional factors. Summary of the Invention

[0004] In view of the aforementioned existing problems, the present invention is proposed.

[0005] Therefore, this invention provides a method for preparing a lotus leaf and black tea compound lipid-lowering substitute tea, which solves the problem that existing technologies, lacking precise spatiotemporal control, cannot induce lotus leaf alkaloids and theabrownins to form a stable nanoscale self-assembled inclusion structure, thus limiting the synergistic lipid-lowering effect of active ingredients and resulting in a bitter taste.

[0006] To solve the above-mentioned technical problems, the present invention provides the following technical solution: In a first aspect, the present invention provides a lotus leaf and black tea compound lipid-lowering substitute tea, which includes enzymatically hydrolyzed lotus leaf powder and highly active black tea base material, which are compounded and then subjected to directional fermentation under the coupled conditions of ultrasonic assistance and segmented temperature and humidity control. The substitute tea contains a nanoscale self-assembled inclusion structure spontaneously formed by lotus leaf alkaloids, theabrownins, and tea polysaccharides through non-covalent bonding. The enzymatically hydrolyzed lotus leaf powder is obtained by enzymatic hydrolysis and cell wall breaking of lotus leaf raw materials, and the highly active black tea base is obtained by co-fermentation and domestication of black tea raw materials with specific probiotics.

[0007] Furthermore, a composite bio-enzyme is used to directionally break down the tough cellulose and pectin cell wall barriers of lotus leaves, releasing intracellular active substances. Simultaneously, specific probiotics are used to pre-acclimate the black tea base material, enriching theabrownin precursors. Based on this, the two are combined and placed in a specially designed ultrasonic fermentation field, implementing segmented temperature and humidity control: In the first stage (aerobic oxidation period), the ultrasonic cavitation effect is used to enhance micro-oxygen mass transfer, accelerating the directional polymerization of tea polyphenols into theabrownins; in the second stage (anaerobic assembly period), by switching to a low-temperature anaerobic environment and adjusting the ultrasonic frequency, non-covalent interactions are induced between hydrophobic lotus leaf alkaloids and amphiphilic theabrownin and tea polysaccharide molecules, spontaneously assembling into thermodynamically stable nanoscale self-assembled inclusion structures. This microstructure not only achieves molecular-level fusion of functional factors but also fundamentally changes the form of the active ingredients, transforming them from a free state to an inclusion state.

[0008] As a preferred embodiment of the lotus leaf and black tea compound lipid-lowering substitute tea of ​​the present invention, the mass ratio of the enzymatically hydrolyzed lotus leaf powder to the highly active black tea base is (3-7):(7-3), wherein the mass ratio is 5:5 or 4:6.

[0009] Furthermore, the optimal mass ratio of enzymatically hydrolyzed lotus leaf powder to highly active dark tea base material was established as (3-7):(7-3), with 5:5 or 4:6 being the preferred core implementation ratio. This ratio range is not a simple quantitative addition, but rather a precise match between the release kinetics of the active ingredients and the metabolic needs of the microorganisms: if the lotus leaf ratio is too high, excessive alkaloids may inhibit the metabolic activity of the dominant bacteria in dark tea (such as *Aspergillus cristatus*), hindering the formation of theaflavins; if the dark tea ratio is too high, it cannot provide sufficient lotus leaf alkaloids to form an effective synergistic lipid-lowering concentration. The preferred 5:5 or 4:6 ratio can construct a microecological balance system during fermentation, ensuring both the full release and retention of lotus leaf alkaloids and providing sufficient substrate and a suitable acid-base environment for the continuous synthesis of theaflavins. This achieves a perfect balance between the mellow taste of dark tea and the delicate aroma of lotus leaf in macroscopic flavor, and a synergistic effect threshold of 1+1>2 in microscopic efficacy.

[0010] As a preferred embodiment of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, wherein: the particle size distribution range of the nanoscale self-assembled inclusion structure is 50nm~300nm, and the average particle size is 100nm~200nm. In the inclusion structure, the encapsulation rate of lotus leaf alkaloid is 75%~90%, and the content of theabrownin in the substitute tea is ≥15wt%, with a retention rate of lotus leaf alkaloid ≥80%.

[0011] Furthermore, dynamic light scattering (DLS) and transmission electron microscopy (TEM) characterization confirmed that the product contains uniformly distributed spherical or ellipsoidal nanocomplexes with particle sizes ranging from 50 nm to 300 nm (average particle size 100 nm to 200 nm). This nanostructure exhibits extremely high encapsulation efficiency, with the lotus leaf alkaloid encapsulation rate consistently between 75% and 90%. This means that the vast majority of the easily oxidized and bitter-tasting lotus leaf alkaloid is tightly encapsulated within a hydrophilic shell formed by theaflavins / tea polysaccharides, effectively preventing degradation and loss during processing and storage, resulting in a lotus leaf alkaloid retention rate of ≥80% in the final product. Simultaneously, thanks to the enhanced targeted fermentation process, the content of the functional core component, theaflavins, in the product is significantly increased and stabilized at ≥15 wt%. This combination of high encapsulation efficiency and high active ingredient content not only ensures the product's stability in a simulated gastric juice environment (reducing gastric irritation) but also achieves targeted sustained release in the intestinal environment, significantly improving bioavailability and lipid-lowering efficacy.

[0012] As a preferred embodiment of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, the compound biological enzyme includes cellulase and pectinase in a mass ratio of (1.5-2.5):1, and the specific probiotics include Eurotium cristatum and Lactobacillus plantarum in an inoculation volume ratio of 2-4):1.

[0013] Furthermore, in the lotus leaf pretreatment stage, a composite enzyme system was selected, consisting of cellulase and pectinase at a mass ratio of (1.5-2.5):1. This ratio produces a synergistic hydrolysis effect, maximizing the destruction of the waxy layer and middle cell wall of the lotus leaf epidermis while avoiding excessive enzymatic hydrolysis that could damage the structure of active ingredients. In the black tea base material domestication stage, an innovative co-culture system was constructed using *Eurotium cristatum* and *Lactobacillus plantarum* at an inoculation volume ratio of (2-4):1. *Eurotium cristatum*, as a key microorganism for black tea flowering, is mainly responsible for secreting extracellular oxidases to promote the conversion of tea polyphenols into theaflavins; while *Lactobacillus plantarum* rapidly lowers the environmental pH by producing acid, inhibiting the growth of other microorganisms and providing necessary growth factors for *Eurotium cristatum*. Simultaneously, the extracellular polysaccharides produced by its metabolism can act as natural emulsifiers to participate in the stability of subsequent nano-inclusion structures. The establishment of this fungal-bacterial symbiotic system provides an ideal biochemical microenvironment for the nano-self-assembly process.

[0014] Secondly, this invention provides a method for preparing a lotus leaf and dark tea compound lipid-lowering substitute tea, comprising, As a preferred embodiment of the preparation method of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, wherein: S1, after selecting lotus leaf raw material and crushing it, enzymatic hydrolysis is performed using compound biological enzymes, and after enzyme inactivation and drying, enzymatic hydrolyzed lotus leaf powder is obtained; black tea raw material is selected, inoculated with specific probiotics for solid-state pre-fermentation and domestication, and after drying, a highly active black tea base material is obtained. S2. Mix the enzymatically hydrolyzed lotus leaf powder obtained in step S1 with the high-activity black tea base material in a predetermined ratio, add sterile water to adjust the moisture content of the material to 50%, and obtain the compound material. S3. Place the compound material in a fermentation container, control the temperature at 30°C and the relative humidity at 80%~90%, introduce a trace amount of oxygen to maintain the dissolved oxygen concentration at 2~5mg / L, apply ultrasonic treatment with the first parameter, ferment for 20 hours to promote the oxidation and polymerization of tea polyphenols to generate theabrownins. S4. Switch the fermentation environment to an anaerobic state, control the temperature to 20°C, adjust the ultrasonic wave to the second parameter for pulse treatment, and continue fermentation for 12-20 hours to induce lotus leaf alkaloids to form nanoscale self-assembled inclusion structures with theabrownins and tea polysaccharides. S5. The fermented material is dried at a temperature below 50°C until the moisture content is ≤5%, then crushed or granulated and packaged to obtain the lotus leaf black tea compound lipid-lowering substitute tea.

[0015] Furthermore, (S1) the lotus leaf cell wall barrier is broken down by targeted enzymatic hydrolysis using compound biological enzymes to release intracellular active ingredients, while specific probiotics are used to pre-ferment and domesticate the black tea in a solid state to enrich theabrownin precursors, thus obtaining a highly bioactive dual-base material; then (S2) the two are compounded in an optimized ratio and the moisture content is precisely controlled to 50% to construct a solid-liquid microenvironment suitable for mass transfer; subsequently, the core segmented coupled fermentation stage begins, first in (S3) under micro-aerobic conditions (dissolved oxygen 2~5mg / L), constant temperature (30℃), and cavitation intensity of the first parameter, ultrasound. Under the action of chemical reaction, the oxidation and polymerization of tea polyphenols are accelerated to generate high content of theabrownins. Then (S4) the environment is switched to a low temperature (20℃) anaerobic environment and a high-power ultrasonic pulse is applied as the second parameter. This induces hydrophobic lotus leaf alkaloids and amphiphilic theabrownins and tea polysaccharides to spontaneously assemble into a thermodynamically stable nanoscale inclusion structure through non-covalent bonds, realizing the molecular-level fusion of functional factors and deastringency and solubilization. Finally (S5) a low temperature drying technology below 50℃ is used to lock the micro-morphology and active ingredients. After pulverization and granulation, a finished substitute tea with both high lipid-lowering effect and mellow taste is obtained.

[0016] As a preferred embodiment of the preparation method of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, in step S1, lotus leaf powder and compound biological enzyme solution are mixed at a material-to-liquid ratio of 1:(6-10), the pH is adjusted to 4.8-5.2, and enzymatic hydrolysis is performed at 40℃-50℃ for 30-50 minutes; then the enzyme is inactivated by heating at 90℃-100℃ for 8-12 minutes.

[0017] Furthermore, the lotus leaf powder and the compound bio-enzyme solution are thoroughly mixed at a material-to-liquid ratio of 1:(6-10). This ratio ensures the effective contact area between the enzyme molecules and the substrate, avoiding uneven enzymatic hydrolysis caused by excessively high local concentrations. The pH value of the reaction system is strictly locked in the weakly acidic range of 4.8~5.2, which is the highest activity window for the synergistic effect of cellulase and pectinase, maximizing the hydrolysis of the cellulose network structure and pectin middle layer in the cell wall. Enzymatic hydrolysis is carried out at a mild temperature of 40℃~50℃ for 30~50 minutes, which can ensure the rate of enzymatic reaction and prevent the thermal degradation of heat-sensitive lotus leaf alkaloids and flavonoids. Immediately afterwards, a high-temperature instantaneous enzyme inactivation treatment at 90℃~100℃ is carried out for 8~12 minutes to completely inactivate the enzyme activity to terminate the reaction, lock in the released active ingredients and generated oligosaccharides, thereby obtaining enzymatically hydrolyzed lotus leaf powder with high dissolution rate and low coldness.

[0018] As a preferred embodiment of the preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea of ​​the present invention, in step S1, the dark tea raw material is moistened to a moisture content of 45%~55%, and a specific probiotic liquid after activation is sprayed on it. The inoculation amount is 2%~4% of the mass of the dark tea raw material. It is fermented in the dark for 3~5 days at 30℃~34℃ and relative humidity of 80%~90% until golden yellow spores appear on the surface of the tea leaves and the oxidation rate of tea polyphenols reaches 25%~35%.

[0019] Furthermore, the raw materials of dark tea are moistened to a moisture content of 45%–55% to provide the necessary free water for microbial metabolism. A specific probiotic solution (a complex of *Aspergillus cristatus* and *Lactobacillus plantarum*) at an inoculation rate of 2%–4% is sprayed on top, and solid-state pre-fermentation is carried out for 3–5 days in a light-proof environment at 30℃–34℃ and relative humidity of 80%–90%. The key control point at this stage is monitoring the formation of golden-yellow spores (*Aspozos cristatus* ascospores) on the surface of the tea leaves and the oxidation rate of tea polyphenols (controlled at 25%–35%). This indicates that the dominant bacterial group has successfully colonized and initiated the initial oxidation and polymerization of tea polyphenols, generating abundant theabrownin precursors. Simultaneously, the acid-producing effect of lactic acid bacteria regulates the substrate pH, inhibiting the growth of other bacteria, and providing a highly active dark tea base material with high enzyme activity and suitable pH for subsequent deep coupling fermentation with lotus leaf components.

[0020] As a preferred embodiment of the preparation method of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, in step S3, the ultrasonic frequency is 25~30kHz, the power density is 0.3~0.5W / mL, and a pulse mode of 2 seconds working and 3 seconds intermittent is adopted. The flow rate of the trace oxygen is 0.5~1.5L / min to ensure that the fermentation system is in a micro-oxygen state.

[0021] Furthermore, a micro-oxygen environment-ultrasonic cavitation dual-field coupling strategy was employed to accelerate the biosynthesis of theaflavins. The fermentation temperature was kept constant at 30℃, with a high humidity of 80%–90%. A trace amount of oxygen was introduced at a flow rate of 0.5–1.5 L / min, precisely controlling the dissolved oxygen concentration within the micro-oxygen range of 2–5 mg / L. This is the optimal oxygen partial pressure for the oxidative condensation of tea polyphenols catalyzed by polyphenol oxidase, avoiding both excessive browning caused by strong oxidation and fermentation stagnation due to hypoxia. Simultaneously, intermittent ultrasound (2 seconds of operation followed by a 3-second interval) at a frequency of 25–30 kHz and a power density of 0.3–0.5 W / mL was applied. The microjets and shock waves generated by the ultrasonic cavitation effect significantly enhanced the mass transfer efficiency of oxygen at the solid-liquid interface, broke the concentration gradient within the fermentation pile, and disrupted some hyphae to release intracellular enzymes, thereby efficiently promoting the directional conversion and polymerization of tea polyphenols into theaflavins.

[0022] In a preferred embodiment of the preparation method of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, in step S4, the anaerobic state is achieved by filling the fermentation container with nitrogen to replace the air, so that the dissolved oxygen concentration is reduced to below 0.5 mg / L; The ultrasonic frequency is 35~45kHz, the power density is 0.5~0.7W / mL, and a pulse mode of 5 seconds working and 5 seconds intermittent is used to induce molecular self-assembly by utilizing the cavitation effect of low-frequency high-power ultrasound.

[0023] Furthermore, high-purity nitrogen is introduced into the fermentation vessel to replace the air, rapidly reducing the dissolved oxygen concentration to below 0.5 mg / L, creating a strictly anaerobic state and cutting off the oxidation reaction pathway. Simultaneously, the temperature is lowered to 20°C to reduce molecular thermal motion, which is conducive to the formation of weak interaction forces. Then, the ultrasonic parameters are adjusted to a low-frequency, high-power mode (frequency 35~45kHz, power density 0.5~0.7W / mL), using a pulse mode of 5 seconds working and 5 seconds intermittent. The stronger cavitation mechanical force induces the hydrophobic lotus leaf alkaloid molecules and the amphiphilic theaflavins and tea polysaccharide molecules to undergo conformational rearrangement. This allows them to spontaneously and orderly arrange themselves around the lotus leaf alkaloid core through hydrogen bonding, stacking, and hydrophobic interactions, forming a uniform particle size and dense nanoscale self-assembled inclusion structure. This achieves the stable encapsulation of functional factors and effective masking of bitterness.

[0024] As a preferred embodiment of the preparation method of the lotus leaf black tea compound lipid-lowering substitute tea of ​​the present invention, wherein: in step S5, the low-temperature drying adopts vacuum freeze drying or microwave vacuum drying process. If vacuum freeze drying is used, the pre-freezing temperature is -35℃ to -45℃, the vacuum degree is 0.05 to 0.15 mbar, the sublimation drying time is 18 to 24 hours, and the desorption drying temperature is 25℃ to 35℃. If microwave vacuum drying is used, the core temperature of the material should always be controlled to not exceed 45℃. Furthermore, advanced low-temperature dehydration technology is employed to fully preserve the heat-sensitive active ingredients and fragile nanostructures. If a vacuum freeze-drying process is used, the material is first pre-frozen to -35℃ to -45℃ to completely solidify the moisture, followed by sublimation drying at a high vacuum of 0.05~0.15 mbar for 18~24 hours to remove most of the free water. Finally, desorption drying at 25℃~35℃ removes bound water. This entire process avoids the damage to the nanostructure caused by the surface tension of liquid water. If a microwave vacuum drying process is used, an intelligent temperature control system strictly limits the material's core temperature to no more than 45℃, utilizing the selective heating characteristics of microwaves to rapidly evaporate moisture under low pressure. Both processes ensure that the final product has a moisture content of ≤5%, while maximizing the retention of theaflavins and lotus leaf alkaloids, preventing high temperatures from causing nanostructure disintegration or oxidative degradation of active ingredients.

[0025] The beneficial effects of this invention are as follows: Through a dual pretreatment step involving the targeted enzymatic hydrolysis of lotus leaf cell walls by compound bio-enzymes and co-fermentation with specific probiotics to domesticate dark tea, the efficient release of active ingredients and enrichment of precursor substances in the raw materials are achieved, increasing the bioavailability of lotus leaf alkaloids, flavonoids, and theabrownin precursors while reducing the cold and irritating properties of lotus leaves. Furthermore, through a segmented temperature and humidity-coupled fermentation step involving ultrasound-assisted oxidation polymerization under a microaerobic environment and ultrasound-induced molecular self-assembly under an anaerobic low-temperature environment, the efficient and targeted conversion of tea polyphenols to theabrownins and the improvement of hydrophobic lotus leaf... Lotus leaf alkaloids spontaneously form a stable nanoscale self-assembled inclusion structure with amphiphilic theabrownins and tea polysaccharides through non-covalent bonds. This unique microstructure not only effectively masks the bitter taste of lotus leaf alkaloids and improves the mellowness of the tea soup, but also protects the stability of heat-sensitive active ingredients during processing and digestion through the nano-encapsulation effect. This significantly enhances the intestinal-targeted sustained release of functional factors and the synergistic lipid-lowering effect. Finally, combined with a low-temperature drying process, this microstructure and high content of active ingredients are locked in, resulting in a functional substitute tea with excellent lipid-lowering performance and superior sensory quality. Attached Figure Description

[0026] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the following description of the embodiments will be briefly introduced. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0027] Figure 1 Flowchart of the preparation method for lotus leaf black tea compound lipid-lowering substitute tea. Detailed Implementation

[0028] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings.

[0029] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0030] Secondly, the term "one embodiment" or "example" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the invention. The appearance of an embodiment in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that mutually excludes other embodiments.

[0031] Example 1, Reference Figure 1This is the first embodiment of the present invention, which provides a lotus leaf and black tea compound lipid-lowering substitute tea and its preparation method, including the following steps: Raw material pretreatment (S1): Lotus leaf processing: After frost, dry lotus leaves are pulverized and passed through a 60-mesh sieve. A compound enzyme solution (cellulase: pectinase = 2:1) is added at a material-to-liquid ratio of 1:8, and the pH is adjusted to 5.0. Enzymatic hydrolysis is carried out at 45℃ for 40 minutes. Subsequently, the enzyme is inactivated by heating at 95℃ for 10 minutes, and the leaves are dried at low temperature to obtain enzymatically hydrolyzed lotus leaf powder.

[0032] Dark tea processing: Moisten Anhua dark tea leaves to a moisture content of 50%. Spray with activated bacterial solution (Eurotium cristatum: Lactobacillus plantarum volume ratio = 3:1), with an inoculation amount of 3%. Ferment in the dark for 4 days at 32℃ and 85% humidity until the tea leaves show golden yellow spores and the tea polyphenol oxidation rate is about 30%. Dry to obtain highly active dark tea base material.

[0033] Compounding and wetting (S2): Mix the enzymatically hydrolyzed lotus leaf powder and the highly active dark tea base material at a mass ratio of 5:5 until homogeneous. Add sterile water to adjust the final moisture content of the material to 50%.

[0034] First stage fermentation (S3-aerobic oxidation): Place in an ultrasonic fermenter, maintain a temperature of 30°C and a humidity of 85%. Introduce sterile air to control dissolved oxygen (DO) at 3.5 mg / L (flow rate approximately 1.0 L / min).

[0035] Apply ultrasound: frequency 28kHz, power density 0.4W / mL, mode "working 2s / intermittent 3s".

[0036] Continue fermentation for 20 hours.

[0037] Second stage fermentation (S4-anaerobic self-assembly): Replace the air with nitrogen to reduce the dissolved oxygen (DO) level to below 0.3 mg / L. Cool down to 20°C.

[0038] Adjust the ultrasound settings: frequency 40kHz, power density 0.6W / mL, mode "5s on / 5s off".

[0039] Continue fermentation for 16 hours.

[0040] Drying and finished product (S5): Vacuum freeze drying was employed: pre-freezing at -40℃, vacuum degree of 0.1mbar, sublimation drying for 20 hours, and desorption drying at 30℃.

[0041] It is crushed through an 80-mesh sieve and then packaged with nitrogen.

[0042] Expected results: The product is dark brown, without any astringent taste, and has a mellow flavor. Testing shows that theabrownin content is ≥18%, lotus leaf alkaloid retention rate is ≥85%, the average particle size of the nano-inclusion complex is approximately 150nm, and the encapsulation rate is approximately 82%.

[0043] Example 2, Reference Figure 1 This is the first embodiment of the present invention, which provides a lotus leaf and black tea compound lipid-lowering substitute tea and its preparation method, including the following steps: A flavor profile emphasizing dark tea (with adjusted proportions) Raw material pretreatment (S1): The enzymatic hydrolysis conditions were the same as in Example 1 (pH 4.8, 40℃, 50 min).

[0044] The acclimatization conditions for black tea were the same as in Example 1 (fermentation for 3 days, oxidation rate of 25%).

[0045] Compounding and wetting (S2): The enzymatically hydrolyzed lotus leaf powder and highly active dark tea base were mixed at a mass ratio of 4:6. The moisture content was adjusted to 50%.

[0046] First stage fermentation (S3): Temperature controlled at 30℃, DO controlled at 2.5mg / L.

[0047] Ultrasound parameters: frequency 25kHz, power density 0.3W / mL, “working 2s / intermittent 3s”.

[0048] Ferment for 20 hours.

[0049] Second stage fermentation (S4): Anaerobic environment (DO<0.5mg / L), cool to 20℃.

[0050] Ultrasound parameters: frequency 35kHz, power density 0.5W / mL, “5s working / 5s resting”.

[0051] Ferment for 12 hours.

[0052] Drying and finished product (S5): Microwave vacuum drying is used: the center temperature of the material is controlled to not exceed 45℃, and the moisture content is dried to ≤5%.

[0053] Expected results: The dark tea will have a richer, more mellow aroma, and its lipid-lowering effect will be slightly lower than in Example 1, but its flavor will be fuller. Theabrownin content will be ≥16%, and the lotus leaf alkaloid retention rate will be ≥80%.

[0054] Example 3, Reference Figure 1 This is the first embodiment of the present invention, which provides a lotus leaf and black tea compound lipid-lowering substitute tea and its preparation method, including the following steps: A scheme emphasizing the delicate fragrance of lotus leaves (with adjusted proportions) Raw material pretreatment (S1): The enzymatic hydrolysis conditions were the same as in Example 1 (pH 5.2, 50℃, 30 min).

[0055] The acclimatization conditions for black tea were the same as in Example 1 (fermentation for 5 days, oxidation rate of 35%).

[0056] Compounding and wetting (S2): The enzymatically hydrolyzed lotus leaf powder and highly active dark tea base were mixed at a mass ratio of 6:4 (Note: within the scope of the claims). The moisture content was adjusted to 50%.

[0057] First stage fermentation (S3): Temperature controlled at 30℃, DO controlled at 4.5mg / L.

[0058] Ultrasound parameters: frequency 30kHz, power density 0.5W / mL, “working 2s / intermittent 3s”.

[0059] Ferment for 20 hours.

[0060] Second stage fermentation (S4): An anaerobic environment was created, and the temperature was lowered to 20°C.

[0061] Ultrasound parameters: frequency 45kHz, power density 0.7W / mL, “5s working / 5s resting”.

[0062] Ferment for 20 hours.

[0063] Drying and finished product (S5): Vacuum freeze drying was used (same as in Example 1).

[0064] Expected results: Distinct lotus leaf aroma, extremely low bitterness (due to good inclusion effect). Theabrownin content ≥15.5%, lotus leaf alkaloid retention rate ≥82%.

[0065] Comparative Example 1: Enzyme Deprivation and Microbial Acclimation (Traditional Physical Mixing) Differences: S1: Lotus leaves are only crushed and not enzymatically hydrolyzed; black tea is not inoculated with microbial cultures for fermentation, and commercially available finished black tea is used directly.

[0066] S2-S5: The process parameters are exactly the same as in Example 1.

[0067] Results analysis: Due to the lack of enzymatic hydrolysis to break down the cell walls, the dissolution rate of lotus leaf alkaloids is low, and the large-molecule polysaccharides are not converted, resulting in a strong cooling sensation in the product, which may easily cause stomach discomfort. Because the dark tea has not undergone pre-acclimation, there is insufficient theabrownin precursor, and the final product contains only about 10% theabrownins. Due to the lack of specific bioactive interfaces, very few nano-inclusion structures are formed, resulting in a noticeable astringent taste, and the retention rate of lotus leaf alkaloids is less than 60%.

[0068] Comparative Example 2: Deletion of segmented coupled fermentation (single-condition fermentation) Differences: S1-S2: Same as Example 1.

[0069] S3-S4 combined: Fermentation was carried out at 30℃ under micro-aerobic (DO3.5mg / L) conditions for 36 hours without anaerobic switching or cooling. The ultrasonic parameters were kept at the first parameter (28kHz, 0.4W / mL) throughout the process.

[0070] S5: Same as Example 1.

[0071] Results analysis: Because the entire process takes place in an aerobic oxidation environment, tea polyphenols continuously oxidize and polymerize. However, lacking the induction of an anaerobic, low-temperature phase, lotus leaf alkaloids cannot form a stable non-covalent inclusion structure with theaflavins. Testing revealed that lotus leaf alkaloids in the product exist primarily in a free state, making them highly susceptible to oxidative degradation, with a retention rate of only about 55%. The product has a bitter taste, and in vitro simulated digestion experiments showed that it is released too quickly in gastric juice.

[0072] Comparative Example 3: Lack of ultrasound assistance (conventional static fermentation) Differences: S1-S2: Same as Example 1.

[0073] S3-S4: Temperature and gas control are the same as in Example 1, but ultrasonic waves are not activated; natural diffusion and mechanical stirring are used instead.

[0074] S5: Same as Example 1.

[0075] Results analysis: The lack of enhanced ultrasonic cavitation results in low oxygen mass transfer efficiency, leading to uneven fermentation and a significant decrease in the production of theaflavins (approximately 12%). Simultaneously, the lack of ultrasonic mechanical induction makes it difficult for molecules to overcome energy barriers and self-assemble, resulting in aggregates with a wide and unstable particle size distribution (500 nm) and an encapsulation rate below 40%. The product has a dull color, a muffled aroma, and an insignificant lipid-lowering synergistic effect.

[0076] In summary, this invention achieves efficient release of active ingredients and enrichment of precursor substances in raw materials through a dual pretreatment process involving the targeted enzymatic hydrolysis of lotus leaf cell walls by compound bio-enzymes and co-fermentation with specific probiotics to domesticate dark tea. This enhances the bioavailability of lotus leaf alkaloids, flavonoids, and theabrownin precursors while reducing the cold and irritating properties of lotus leaves. Furthermore, through a segmented temperature and humidity-coupled fermentation process involving ultrasound-assisted oxidation polymerization under microaerobic conditions and ultrasound-induced molecular self-assembly under anaerobic low-temperature conditions, this invention achieves efficient and targeted conversion of tea polyphenols to theabrownins and the processing of hydrophobic lotus leaves. The alkali spontaneously forms a stable nanoscale self-assembled inclusion structure with the amphiphilic theaflavins and tea polysaccharides through non-covalent bonds. This unique microstructure not only effectively masks the bitter taste of lotus leaf alkaloids and improves the mellowness of the tea soup, but also protects the stability of heat-sensitive active ingredients during processing and digestion through the nano-encapsulation effect. This achieves a significant enhancement of the intestinal-targeted sustained release of functional factors and synergistic lipid-lowering effects. Finally, combined with a low-temperature drying process, this microstructure and high content of active ingredients are locked in, resulting in a functional substitute tea with excellent lipid-lowering performance and superior sensory quality.

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

Claims

1. A lotus leaf and dark tea compound lipid-lowering substitute tea, characterized in that: This includes enzymatically hydrolyzed lotus leaf powder and highly active black tea base material, which are compounded and then subjected to directional fermentation under coupled conditions of ultrasonic assistance and segmented temperature and humidity control. The substitute tea contains a nanoscale self-assembled inclusion structure spontaneously formed by lotus leaf alkaloids, theabrownins, and tea polysaccharides through non-covalent bonding. The enzymatically hydrolyzed lotus leaf powder is obtained by enzymatic hydrolysis and cell wall breaking of lotus leaf raw materials, and the highly active black tea base is obtained by co-fermentation and domestication of black tea raw materials with specific probiotics.

2. The lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 1, characterized in that: The mass ratio of the enzymatically hydrolyzed lotus leaf powder to the highly active black tea base is (3-7):(7-3), where the mass ratio is 5:5 or 4:

6.

3. The lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 2, characterized in that: The particle size distribution range of the nanoscale self-assembled inclusion structure is 50nm~300nm, and the average particle size is 100nm~200nm; In the inclusion structure, the encapsulation rate of lotus leaf alkaloid is 75%~90%, and the content of theabrownin in the substitute tea is ≥15wt%, with a retention rate of lotus leaf alkaloid ≥80%.

4. The lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 3, characterized in that: The composite bioenzyme includes cellulase and pectinase in a mass ratio of (1.5-2.5):1, and the specific probiotics include Eurotium cristatum and Lactobacillus plantarum in an inoculation volume ratio of 2-4):

1.

5. A method for preparing a lotus leaf dark tea compound lipid-lowering substitute tea, based on the lotus leaf dark tea compound lipid-lowering substitute tea according to any one of claims 1 to 4, characterized in that: include, S1. Select lotus leaf raw materials, crush them, and then use compound biological enzymes to perform enzymatic hydrolysis. After enzyme inactivation and drying, enzymatic hydrolyzed lotus leaf powder is obtained. Select black tea raw materials, inoculate them with specific probiotics for solid-state pre-fermentation and domestication, and then dry them to obtain highly active black tea base material. S2. Mix the enzymatically hydrolyzed lotus leaf powder obtained in step S1 with the high-activity black tea base material in a predetermined ratio, add sterile water to adjust the moisture content of the material to 50%, and obtain the compound material. S3. Place the compound material in a fermentation container, control the temperature at 30°C and the relative humidity at 80%~90%, introduce a trace amount of oxygen to maintain the dissolved oxygen concentration at 2~5mg / L, apply ultrasonic treatment with the first parameter, ferment for 20 hours to promote the oxidation and polymerization of tea polyphenols to generate theabrownins. S4. Switch the fermentation environment to an anaerobic state, control the temperature to 20°C, adjust the ultrasonic wave to the second parameter for pulse treatment, and continue fermentation for 12-20 hours to induce lotus leaf alkaloids to form nanoscale self-assembled inclusion structures with theabrownins and tea polysaccharides. S5. The fermented material is dried at a temperature below 50°C until the moisture content is ≤5%, then crushed or granulated and packaged to obtain the lotus leaf black tea compound lipid-lowering substitute tea.

6. The preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 5, characterized in that: In step S1, lotus leaf powder and compound biological enzyme solution are mixed at a material-to-liquid ratio of 1:(6-10), the pH is adjusted to 4.8-5.2, and enzymatic hydrolysis is carried out at 40℃-50℃ for 30-50 minutes; then the enzyme is inactivated by heating at 90℃-100℃ for 8-12 minutes.

7. The preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 5, characterized in that: In step S1, the black tea raw material is moistened to a moisture content of 45% to 55%, and a specific probiotic liquid after activation is sprayed on it. The inoculation amount is 2% to 4% of the mass of the black tea raw material. It is fermented in the dark for 3 to 5 days at 30℃ to 34℃ and relative humidity of 80% to 90% until golden yellow spores appear on the surface of the tea leaves and the oxidation rate of tea polyphenols reaches 25% to 35%.

8. The preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 5, characterized in that: In step S3, the ultrasonic frequency is 25~30kHz, the power density is 0.3~0.5W / mL, and a pulse mode of 2 seconds working and 3 seconds intermittent is adopted. The flow rate of the trace oxygen is 0.5~1.5L / min to ensure that the fermentation system is in a micro-oxygen state.

9. The preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 5, characterized in that: In step S4, the anaerobic state is achieved by introducing nitrogen into the fermentation vessel to replace the air, thereby reducing the dissolved oxygen concentration to below 0.5 mg / L; The ultrasonic frequency is 35~45kHz, the power density is 0.5~0.7W / mL, and a pulse mode of 5 seconds working and 5 seconds intermittent is used to induce molecular self-assembly by utilizing the cavitation effect of low-frequency high-power ultrasound.

10. The preparation method of the lotus leaf dark tea compound lipid-lowering substitute tea as described in claim 5, characterized in that: In step S5, the low-temperature drying is performed using vacuum freeze-drying or microwave vacuum drying processes. If vacuum freeze drying is used, the pre-freezing temperature is -35℃ to -45℃, the vacuum degree is 0.05 to 0.15 mbar, the sublimation drying time is 18 to 24 hours, and the desorption drying temperature is 25℃ to 35℃. If microwave vacuum drying is used, the core temperature of the material should always be controlled to not exceed 45℃.

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