Functional tea extract as well as preparation method and application thereof

Abstract

The invention discloses a preparation method and application of a functional tea extract, and aims to solve the problems that active ingredients are easy to oxidize and degrade and low in purity in existing tea leaf processing. Enzymatic browning is inhibited through liquid nitrogen quick-freezing pretreatment, the mass transfer efficiency is enhanced in combination with dynamic ultrasonic-assisted extraction, efficient enrichment is achieved by adopting a multi-stage membrane separation or resin purification technology, and the retention rate and purity of catechin substances (especially epigallocatechin gallate EGCG) are remarkably improved. According to the method, thermal degradation and oxidation loss are effectively reduced, and the obtained extract has excellent antioxidant and antibacterial properties and can be widely applied to the fields of functional food, health care products and the like.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of natural product deep processing and functional ingredient development, in particular to a method for preparing high-purity and high-activity tea extract and its application in the fields of antioxidant, antibacterial and the like. BACKGROUND

[0002] Tea is a natural drink widely consumed worldwide, and its health value mainly comes from the polyphenolic active substances rich in tea leaves, especially catechin ingredients. Among them, epigallocatechin gallate (EGCG) has become a research hotspot and application focus in the fields of functional food, health products and cosmetics, etc. due to its strong antioxidant, antibacterial, anti-inflammatory and other biological activities.

[0003] At present, the conventional methods for extracting tea polyphenols and EGCG from tea leaves mainly include hot water extraction, organic solvent extraction and ultrasonic-assisted extraction, etc. However, these traditional processes still have obvious limitations in practical application: first, the hot water extraction method has a high operating temperature, and long-term heat action can easily cause the oxidative degradation and isomerization of heat-sensitive components such as catechins, resulting in a significant decrease in biological activity; second, although the organic solvent (such as acetone, ethanol, ethyl acetate, etc.) extraction method improves the extraction efficiency to some extent, the residual organic solvent is difficult to completely remove, which poses safety and environmental hazards, and the selective enrichment effect of EGCG is limited, and the product purity is not high; third, although the conventional ultrasonic-assisted extraction method can strengthen mass transfer through cavitation effect, it lacks temperature control and inert gas protection in the whole process, and the active ingredients are still easily damaged by heat and oxygen during the extraction process, and the subsequent purification of the extraction liquid is insufficient, resulting in high impurity content.

[0004] Therefore, the tea extract prepared by the prior art generally has the problems of low retention rate of active ingredients, low purity of EGCG, high residual amount of impurities (such as caffeine, pigments, polysaccharides, etc.), poor product quality stability, etc., which is difficult to meet the demand for high-purity and high-activity tea extract for high-end functional products. Developing a green and fine preparation process that can maximize the retention of active ingredients, efficiently enrich target monomers, and avoid solvent pollution and thermal damage has become a technical problem to be solved in the field. SUMMARY

[0005] In order to obtain a functional tea extract with higher content of active substances, the purpose of the present application is to provide a functional tea extract and a preparation method and application thereof.

[0006] In a first aspect, the present application provides a preparation method of a functional tea extract, which adopts the following technical solution: (1) Raw material pretreatment: after removing impurities from fresh tea leaves, liquid nitrogen quick freezing treatment is performed, and the tea leaves are stored in a low-temperature environment, and thawed to room temperature before use; (2) Cell disruption and homogenate preparation: the thawed tea leaves are mixed with pre-cooled purified water or citric acid-sodium citrate buffer with pH=4.0-6.0, and are subjected to ultrafine pulverization under cooling conditions to obtain a tea slurry suspension; (3) Dynamic ultrasonic-assisted extraction: the tea slurry suspension is subjected to multi-frequency ultrasonic extraction under nitrogen protection and micro-positive pressure conditions to obtain a tea slurry mixture; (4) Purification: a membrane separation system is used to purify the tea slurry mixture after ultrasonic extraction to obtain a clear tea concentrate; (5) Drying: the tea concentrate is subjected to stage-by-stage drying, including vacuum belt drying and freeze-sublimation drying, to obtain a functional tea extract; The functional tea extract has tea polyphenols as the main active ingredient, and the content of catechin substances in the tea polyphenols is the highest; the catechin substances contain epigallocatechin gallate (EGCG), and EGCG is the monomer component with the highest content in the catechin substances.

[0007] By using the above technical solution, the retention rate and purity of catechin substances, especially epigallocatechin gallate EGCG, are significantly improved.

[0008] Optionally, the power of the multi-frequency ultrasonic extraction is set to 5-10 w, and the working mode is an intermittent working mode. The intermittent working mode is ultrasonic for 20-40 s and pause for 10-20 s.

[0009] By using the above technical solution, intermittent ultrasonic action can effectively avoid local overheating, reduce the degradation of heat-sensitive components, improve the mass transfer efficiency, and help improve the extraction rate and stability of target components.

[0010] Optionally, 0.01%-0.1%wt of L-ascorbyl palmitate is added during ultrasonic extraction in step (3).

[0011] By using the above technical solution, L-ascorbyl palmitate as a fat-soluble antioxidant can further inhibit the oxidation of catechin substances during the extraction process, and significantly improve the retention rate of tea polyphenols.

[0012] Optionally, the membrane separation system in step (4) is a three-stage membrane series system, including: Primary microfiltration: ceramic microfiltration membranes with a pore size of 0.1-0.5 μm are used to remove suspended solids and macromolecular proteins; Secondary ultrafiltration: ultrafiltration membranes with a molecular weight cut-off of 0.5-2 kDa are used for concentration; Tertiary nanofiltration: nanofiltration membranes with a molecular weight cut-off of 200-500 Da are used for desalination and removal of small molecular impurities.

[0013] By adopting the technical scheme, the three-stage membrane system can gradually remove impurities of different particle sizes, realize efficient enrichment and purification of target components, and significantly improve the purity of active substances such as epigallocatechin gallate (EGCG).

[0014] Optionally, the secondary ultrafiltration step in step (4) adopts a chitosan / cellulose composite modified ultrafiltration membrane.

[0015] By adopting the technical scheme, the composite membrane material has good hydrophilicity and anti-pollution performance, can improve the ultrafiltration flux and the recovery rate of target components, and further improve the purity of the extract.

[0016] Optionally, the tertiary nanofiltration step in step (4) adopts a sodium alginate coated nanofiltration membrane.

[0017] By adopting the technical scheme, the sodium alginate coating has good selectivity and biocompatibility, can effectively desalt and remove small molecule impurities, and improve the purity and safety of the final product.

[0018] Optionally, the purification in step (4) is to adsorb the tea slurry mixture obtained in step (3) by using a weakly polar macroporous adsorption resin, elute and collect target fractions by using water and gradient ethanol solution in sequence, and treat the eluate by using a membrane separation system to obtain a clear tea concentrate.

[0019] By adopting the technical scheme, resin adsorption can target enrichment of high-value components such as EGCG, combined with membrane system refining, which significantly improves the purity and yield of target components.

[0020] In a second aspect, the present application provides a functional tea extract, wherein the total amount of catechins is ≥895 mg / g, the content of epigallocatechin gallate (EGCG) is ≥562 mg / g, the retention rate of tea polyphenols is ≥96.5%, and the content of caffeine is ≤48.2 mg / g.

[0021] By adopting the technical scheme, the extract has the characteristics of high purity and low caffeine, and is suitable for the development of functional products for people sensitive to caffeine.

[0022] Optionally, the functional tea extract has an IC 50 ≤18.5 μg / mL, and a minimum inhibitory concentration (MIC) of Staphylococcus aureus ≤250 μg / mL.

[0023] In a third aspect, the present application provides an application of a functional tea extract, which can be used to prepare functional food, health products, cosmetics or pharmaceutical compositions.

[0024] By adopting the technical scheme, the extract exhibits excellent antioxidant and antibacterial activities, and expands the application potential of the extract in the fields of functional food, health products, cosmetics and the like.

[0025] To sum up, the present application includes at least one of the following beneficial technical effects: 1. By using the pretreatment of liquid nitrogen quick freezing combined with low-temperature crushing and ultrasonic extraction under nitrogen protection, the enzymatic browning and thermal oxidation reaction in the processing process are effectively blocked, and the stability of active ingredients (such as catechin substances) is significantly improved.

[0026] 2. By using a three-stage membrane separation system (microfiltration-ultrafiltration-nanofiltration) or a weakly polar macroporous resin adsorption-elution technology, impurities are accurately removed and target components (such as epigallocatechin gallate EGCG) are enriched, and the purity of the core active substances of the extract is greatly improved.

[0027] 3. Based on the stable retention of high-purity active ingredients, the obtained extract exhibits better performance in antioxidant and antibacterial biological functions, and expands the application potential of the extract in functional products. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 is a schematic diagram of embodiment 8 of the present application.

[0029] The embodiments of the present application will be described in detail below with reference to the examples, but those skilled in the art will understand that the following examples are only used to illustrate the present application and should not be regarded as limiting the scope of the present application. The specific conditions not mentioned in the examples are carried out according to the conventional conditions or the conditions recommended by the manufacturer. The materials or instruments not mentioned by the manufacturer are conventional products that can be purchased on the market.

[0030] The specific embodiments of the present application will be described in detail below. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present application, and are not used to limit the present application.

[0031] Example 1: Tea extract processing method (1) Raw material pretreatment Fresh and complete tea leaves are selected, and old leaves, branches and visible impurities are manually removed, then liquid nitrogen quick freezing treatment (-196℃) is immediately carried out, and the frozen raw materials are stored in a low-temperature environment. Before use, the frozen raw materials are taken out and naturally thawed to room temperature.

[0032] (2) Cell disruption and homogenate preparation The thawed tea leaves were mixed with purified water pre-cooled to 4°C at a mass-volume ratio of 1:8 (g / mL) and injected into a super-micro grinding device equipped with a refrigeration system for wet grinding. The main shaft speed of the grinder was set to 12000 rpm and the grinding was continued for 3 minutes to obtain a uniform tea slurry suspension with a particle size of no more than 5 μm. The entire crushing process was strictly temperature-controlled through a circulating cooling system to ensure that the material temperature was always ≤8°C.

[0033] (3) Dynamic ultrasonic-assisted extraction The obtained tea slurry suspension was transferred to a multi-frequency ultrasonic extraction device with temperature control function, and the power density was set to 6 W / mL. The intermittent working mode was adopted (30 seconds of single ultrasonic action followed by 15 seconds of pause), and the cumulative extraction time was controlled within 20 minutes. Nitrogen was continuously introduced into the system during the extraction process to maintain the reaction environment in a slightly positive pressure state (0.1 MPa).

[0034] (4) Gradient membrane separation and purification A three-stage serial membrane separation system was used to sequentially refine the extract: First-stage treatment: ceramic microfiltration membrane with a pore size of 0.2 μm (item number: CRM-MF) was selected to effectively remove suspended solids and large molecular proteins; Second-stage concentration: selective enrichment was performed by ultrafiltration membrane with a molecular weight cutoff of 1 kDa (item number: BONA-GM-T01) to achieve preliminary concentration of the target product; Third-stage refinement: desalination was performed using nanofiltration membrane with a molecular weight cutoff of 300 Da (item number: HNF40-4040) to simultaneously remove small molecular impurities and residual solvents, and finally a clear concentrated liquid with a transmittance of ≥95% was obtained.

[0035] (5) Stepwise drying and molding A three-stage drying process was performed on the purified concentrated liquid: Pre-drying stage: a vacuum belt dryer was used, the cavity temperature was set to -45°C, the vacuum degree was 0.08 MPa, the liquid was uniformly spread to form a thin layer with a thickness of ≤2 mm, and the water content was reduced to 40%; Main drying stage: the material was subjected to deep dehydration in a freeze-drying equipment under the conditions of -50°C low temperature and <10 Pa high vacuum until the material reached a constant weight; Post-treatment process: after drying, high-purity nitrogen was filled for replacement protection, and after screening to remove lumps, a tea extract powder with moderate apparent density and loose porous texture was obtained.

[0036] Example 2 The difference between this example and Example 1 is that a cross-linked chitosan / cellulose composite ultrafiltration membrane is used for the second-stage concentration in the gradient membrane separation and purification.

[0037] Example 3 The difference between this example and Example 1 is that the third-stage refining in gradient membrane separation and purification uses a biocompatible sodium alginate-coated nanofiltration membrane.

[0038] Example 4 The difference between this example and Example 1 is that in step (2), the thawed tea leaves are mixed with a citric acid-sodium citrate buffer solution and injected into a super-micro pulverizing device equipped with a refrigeration system for wet pulverization.

[0039] Example 5 The difference between this example and Example 1 is that in step (3), 0.05% wt of L-ascorbyl palmitate is added.

[0040] Example 6: (1) Raw material pretreatment: the same as Example 1; (2) Cell disruption and homogenate preparation: the same as Example 1; (3) Dynamic ultrasonic-assisted extraction: the same as Example 1; (4) Resin column purification and membrane system refining: Resin column purification: (a) Resin column preparation: weakly polar macroporous adsorption resin (D101 type) is selected and packed in a glass chromatography column (diameter-height ratio 1:8), and purified water is used for flushing until the effluent is clear for standby; (b) Sample adsorption: the above tea slurry supernatant is pumped into the resin column at a flow rate of 2 BV / h, and the effluent is collected until it is negative by Molish reaction detection; (c) Water washing: the resin column is washed with 2 column volumes of purified water to remove water-soluble sugars and small molecular impurities adsorbed on the surface of the resin; (d) Gradient elution: 30%, 50%, and 70% ethanol solutions are used for stepwise elution, and high-performance liquid chromatography (HPLC) is used for online monitoring of the elution process. The target elution fraction with an EGCG peak area ratio of more than 90% is collected; (e) Resin regeneration: 95% ethanol solution is used to flush the resin until the effluent is colorless, and then purified water is used for flushing until it is neutral for standby;

[0041] Membrane system refining: The eluate containing the target elution fraction is sequentially passed through a three-stage membrane separation system, and the specific method is the same as that in Example 1. Finally, high-purity epigallocatechin gallate (EGCG) enriched liquid is obtained; (5) Stepwise drying and molding The enriched liquid obtained in step (4) was subjected to a three-stage drying process in the same manner as in Example 1, and finally a functional tea extract powder enriched in epigallocatechin gallate (EGCG) was obtained.

[0042] Comparative Example 1: Conventional hot water extraction method Raw material treatment: The same batch of fresh leaves as in Example 1 was taken, and after harvesting, the fresh leaves were dried in a hot air oven at 60°C until the weight was constant, and then mechanically pulverized through a 40-mesh sieve to obtain dry tea powder;

[0043] Extraction: The dry tea powder was added to purified water at 85°C at a mass-to-volume ratio of 1:20 (g / mL), and constant-temperature stirring extraction was performed for 60 minutes;

[0044] Solid-liquid separation: After the extraction was completed, the hot filtrate was collected by coarse filtration with double-layer filter cloth. The filter residue was added to purified water at 85°C at a ratio of 1:10, and repeated extraction was performed for 30 minutes, followed by filtration again;

[0045] Concentration: The two filtrates were combined, and rotary evaporation was performed at 70°C under vacuum at -0.08 MPa to concentrate the filtrate to one-fourth of the original volume;

[0046] Drying: The concentrated liquid was poured into a tray and dried in a blast drying oven at 75°C until the weight was constant. After pulverization, a tea extract powder was obtained.

[0047] Comparative Example 2: Organic solvent extraction method Raw material treatment: The same as in Comparative Example 1, dry tea powder was prepared.

[0048] Solvent extraction: The dry tea powder was added to a 70% (by volume) acetone aqueous solution at a mass-to-volume ratio of 1:15 (g / mL), and stirring extraction was performed three times at 50°C under light-avoiding conditions, each time for 45 minutes;

[0049] Solvent recovery: The extract was combined, and rotary evaporation was performed at 45°C to recover the acetone solvent;

[0050] Desolventization: The remaining aqueous phase was extracted with an equal volume of ethyl acetate three times, and the organic phase was combined. Ethyl acetate was removed again using a rotary evaporator;

[0051] Water-solubilization and drying: The residue was dissolved with a small amount of purified water, and concentration and drying were performed in the manner described in Comparative Example 1 to obtain a tea extract powder.

[0052] Comparative Example 3: Conventional ultrasonic-assisted extraction method Raw material treatment: The same as in Example 1, the fresh leaves were quick-frozen in liquid nitrogen and then thawed;

[0053] Ultrasonic extraction: the thawed fresh leaves were mixed with room temperature pure water at a ratio of 1:10 (g / mL), and placed in a conventional ultrasonic cleaner (rated power 800 W, frequency 40 kHz) for ultrasonic extraction for 30 minutes. No temperature control and atmosphere protection measures were taken during the extraction process; Solid-liquid separation: the mixture after extraction was centrifuged at 4000 rpm for 15 minutes, and the supernatant was taken; Concentration and drying: the supernatant was concentrated by rotary evaporation and air-dried as described in Comparative Example 1 to obtain tea extract powder.

[0054] Example 7

[0055] In this example, the tea leaf extract powders prepared in Examples 1-6 were analyzed for quality, and compared with the traditional method extracts of Comparative Examples 1-3. The catechin content was detected by high performance liquid chromatography, the EGCG content was calculated from the catechin content detection peak graph, the tea polyphenol retention rate was detected and calculated by the Folin phenol method, and the caffeine content was also detected by high performance liquid chromatography. The final results are as follows: Table 1: Analysis of active ingredient content of products of different examples and comparative examples

[0056] Note: The data is expressed as mean ± standard deviation (n=3). Example 5 has different calculation basis due to the addition of antioxidants, and the retention rate exceeds 100%. Example 6 has a retention rate exceeding 100% because it uses targeted enrichment technology.

[0057] In summary, each example (1-6) uses liquid nitrogen quick freezing, low temperature crushing, nitrogen protection ultrasonic, and membrane separation / resin purification, etc. comprehensive means, greatly reducing the oxidation and thermal degradation in the processing process, the content and retention rate of the core active substance (total catechin, epigallocatechin gallate EGCG) are significantly higher than all the comparative examples. Among them, Example 6 targets the enrichment of epigallocatechin gallate (EGCG) by resin adsorption, showing the highest purity of the target component.

[0058] Example 8 Antioxidant capacity evaluation DPPH free radical scavenging capacity Detection method: accurately weigh each sample, dissolve with ethanol and configure into a series of concentration gradients. Take 2 mL of sample solution of different concentrations, add 2 mL of 0.2 mmol / L DPPH ethanol solution, vortex mix, and react at room temperature for 30 minutes in the dark. Measure the absorbance value at 517 nm wavelength, and use the same concentration of vitamin C as a positive control. Calculate the half-inhibitory concentration (IC 50 value) of each sample for DPPH free radical scavenging. 50The lower the value, the stronger the scavenging ability.

[0059] Table 2: Evaluation results of DPPH radical scavenging ability (IC 50 , μg / mL)

[0060] By DPPH method, it is clear that the antioxidant capacity of the extract in the format example is significantly better than all the comparative examples, because in the examples, the retention rate of active ingredients of the extract is higher than that of the comparative examples, so each example also shows strong free radical scavenging ability in the experimental process. Example 5 and Example 6 respectively because of the addition of antioxidants and targeted enrichment of catechins, show the most excellent antioxidant performance in the examples.

[0061] Based on the IC 50 value of 18.5 μg / mL measured in Example 1, the clearance rate curve of the sample prepared in Example 1 at different concentrations was further detected, and Comparative Example 3 was used as a control, Figure 1 The clearance rate of DPPH free radicals of Example 1 and Comparative Example 3 samples with concentration change is shown in the comparative curve.

[0062] From Figure 1 it can be seen that the dose-effect of Comparative Example 3 is significantly right-shifted compared with Example 1 as a whole. When reaching 50% clearance rate, the sample concentration required by Comparative Example 3 is much higher than that required by Example 1. It is explained that to obtain the same antioxidant effect, the sample required by Comparative Example 3 needs to increase by about 75.7%. In the experimental concentration range, the maximum clearance rate plateau of Comparative Example 3 sample is lower. Even if the concentration is increased to 100 μg / mL, its clearance rate is only similar to the effect of Example 1 at 30 μg / mL, and it has always failed to reach the 100% clearance level that Example 1 can achieve at a lower concentration. This significant difference is due to the difference in process between the two. Comparative Example 3 does not perform temperature control and inert gas protection during extraction, resulting in thermal degradation and oxidation of active substances such as catechins, and does not use membrane separation purification steps, so that the impurity content in the extract is high, the proportion of effective components is the lowest, and finally the unit mass of biological activity of Example 1 is significantly better than that of Comparative Example 3.

[0063] Example 9 Anti-bacterial and anti-inflammatory activity verification Detection method: The minimum inhibitory concentration (MIC) was determined by the microdilution broth method (M07-A10) recommended by the Clinical and Laboratory Standards Institute (CLSI). Staphylococcus aureus (ATCC 6538) was used as a representative of gram-positive bacteria, and Escherichia coli (ATCC 8739) was used as a representative of gram-negative bacteria. The sample to be tested was serially diluted in Mueller-Hinton Broth (MHB) medium to prepare a gradient concentration of working solution. Then, the prepared bacterial suspension (final concentration of about 5 x 10 5 CFU / mL) was inoculated and incubated in a constant temperature incubator at 35±2°C for 16-20 hours. The lowest sample concentration without visible bacterial growth was observed as the MIC value of the sample to the tested strain. At the same time, the medium without the sample was set as a positive growth control, and the sample containing medium without bacteria was set as a negative sterile control.

[0064] Table 3: Evaluation results of antibacterial activity (minimum inhibitory concentration MIC, μg / mL) Sample Staphylococcus aureus Escherichia coli Example 1 125 250 Example 2 120 245 Example 3 122 248 Example 4 110 235 Example 5 105 225 Example 6 62.5 125 Comparative Example 1 >500 >500 Comparative Example 2 200 400 Comparative Example 3 250 500 The antibacterial experiment results show that the extracts prepared by each example all exhibit obvious inhibitory effect on gram-positive bacteria (Staphylococcus aureus) and gram-negative bacteria (Escherichia coli), and the antibacterial activity is significantly better than that of the comparative example. The antibacterial activity of the Yongping variety shows a significant negative correlation with the content of epigallocatechin gallate (EGCG), that is, the higher the content of epigallocatechin gallate (EGCG), the lower the MIC value, and the stronger the antibacterial activity. In Example 6, the purity of epigallocatechin gallate (EGCG) in the final product obtained by resin purification is the highest among all examples, so it also exhibits the strongest antibacterial activity. In Comparative Example 1, the active ingredients in the final product are severely damaged due to the hot water extraction method, so there is almost no antibacterial effect.

[0065] Through systematic comparative study of tea leaf extracts prepared by Examples 1-6 and Comparative Examples 1-3 (Tables 1-3), the experimental data show that the composite processing technology used in the present application has significant technical advantages. Specifically: (1) in terms of core active ingredient retention, the total amount of catechins (895.6-1052.4 mg / g) of Examples 1-6 is increased by 150%-298% compared to the traditional hot water extraction method (352.4 mg / g), the content of epigallocatechin gallate (EGCG) (562.8-855.6 mg / g) is increased by 30%-88% compared to the organic solvent extraction method (450.2 mg / g), and the tea polyphenol retention rate (96.5%-100.5%) is all above 95%; (2) in terms of biological activity, the DPPH free radical scavenging capacity of Examples 5 and 6 is 15.2 μg / mL and 12.6 μg / mL, respectively, which is increased by 59%-68% compared to the traditional ultrasonic extraction method (32.5 μg / mL), and the IC 50 values of Examples 5 and 6 and vitamin C (5.2 μg / mL) still differ by an order of magnitude; (3) in terms of antibacterial performance, the MIC value of Example 6 against Staphylococcus aureus is as low as 62.5 μg / mL, which is increased by more than 7 times compared to the hot water extraction method (>500 μg / mL), and the purity of epigallocatechin gallate (EGCG) is positively correlated with the antibacterial activity.

[0066] In summary, through systematic data comparison and functional verification of Examples 1-6 and Comparative Examples 1-3 in Examples 7-9, it is fully shown that the tea leaf extract processing method provided by Examples 1-6 can significantly increase the content and retention rate of various active substances in the final product compared to the traditional process (Comparative Examples 1-3), and greatly improve the biological functions such as antioxidant, free radical scavenging and antibacterial.

[0067] The examples of the specific embodiments are the preferred embodiments of the present application, and are not intended to limit the protection scope of the present application, wherein the same parts are denoted by the same reference numerals. Therefore: any equivalent changes made according to the structure, shape, principle of the present application should be covered within the protection scope of the present application.

Claims

1. A method of preparing a functional tea extract, characterized by, The functional tea extract is prepared by a method comprising the following steps: (1) raw material pretreatment: after removing impurities from fresh tea leaves, liquid nitrogen quick freezing treatment is performed, and the tea leaves are stored in a low temperature environment, and thawed to room temperature before use; (2) cell disruption and homogenate preparation: the thawed tea leaves are mixed with pre-cooled purified water or a citric acid-sodium citrate buffer solution with pH = 4.0-6.0, and are subjected to ultrafine grinding under cooling conditions to obtain a tea slurry suspension; (3) dynamic ultrasonic-assisted extraction: the tea slurry suspension is subjected to multi-frequency ultrasonic extraction under nitrogen protection and micro-positive pressure conditions to obtain a tea slurry mixture; (4) purification: the tea slurry mixture after ultrasonic extraction is subjected to purification by a membrane separation system to obtain a clear tea leaf concentrate; (5) drying: the tea leaf concentrate is subjected to staged drying, including vacuum belt drying and freeze sublimation drying, to obtain the functional tea extract; The functional tea extract takes tea polyphenols as the main active ingredient, and the content of catechin substances in the tea polyphenols is the highest; the catechin substances contain epigallocatechin gallate (EGCG), and EGCG is the monomer component with the highest content in the catechin substances.

2. The method of preparing a functional tea extract according to claim 1, characterized in that: In step (3), the power of the multi-frequency ultrasonic extraction is set to 5-10 W, and the working mode is an intermittent working mode, wherein the intermittent working mode is ultrasonic for 20-40 s and pause for 10-20 s.

3. The method of preparing a functional tea extract according to claim 1, characterized in that, In step (3), 0.01%-0.1% of L-ascorbyl palmitate is added during ultrasonic extraction.

4. The method of preparing a functional tea extract according to claim 1, characterized in that, The membrane separation system in step (4) is a three-stage membrane series system, comprising: primary microfiltration: ceramic microfiltration membranes with a pore size of 0.1-0.5 μm are used to remove suspended solids and macromolecular proteins; secondary ultrafiltration: ultrafiltration membranes with a molecular weight cut-off of 0.5-2 kDa are used for concentration; tertiary nanofiltration: nanofiltration membranes with a molecular weight cut-off of 200-500 Da are used for desalting and removing small molecular impurities.

5. The method of preparing a functional tea extract according to claim 4, characterized in that, The secondary ultrafiltration step in step (4) uses a chitosan / cellulose composite modified ultrafiltration membrane as the ultrafiltration membrane.

6. The method of preparing a functional tea extract according to claim 4, characterized in that, The tertiary nanofiltration step in step (4) uses a sodium alginate coated nanofiltration membrane as the nanofiltration membrane.

7. The method for preparing the functional tea extract according to claim 1, characterized in that, The purification in step (4) uses a weakly polar macroporous adsorption resin to adsorb the tea slurry mixture obtained in step (3), and then water and gradient ethanol solution are used for elution and the target fraction is collected; the eluate is treated by a membrane separation system to obtain a clear tea leaf concentrate.

8. A functional tea extract prepared according to the method of any one of claims 1 to 7, characterized in that, The total amount of catechins in the extract is ≥895 mg / g, the content of epigallocatechin gallate (EGCG) is ≥562 mg / g, the tea polyphenol retention rate is ≥96.5%, and the caffeine content is ≤48.2 mg / g.

9. The functional tea extract according to claim 8, characterized in that, The extract has an IC50 of 18.5 μg / mL 50 ≤ 18.5 μg / mL, minimum inhibitory concentration MIC against Staphylococcus aureus ≤ 250 μg / mL.

10. Use of the functional tea extract of any one of claims 8-9 in the preparation of functional food, health products, cosmetics or pharmaceutical compositions.

Images