Tea leaf fermentation liquor rich in functional components and preparation method of tea leaf fermentation liquor

Abstract

The invention discloses tea leaf fermentation liquor rich in functional components and a preparation method of the tea leaf fermentation liquor, and relates to the technical field of deep processing of tea leaves. The preparation method comprises the following steps: mixing tea leaves with water, and performing enzymolysis to obtain tea leaf zymolyte; culturing the microbial strain; performing microbial inoculation on the tea leaf zymolyte to obtain tea leaf fermentation liquor rich in theanine, theanine and gamma-aminobutyric acid; according to the invention, high-value utilization of the trimmed tea leaves can be realized; after tender shoots and tender leaves of tea trees are picked, most of the tea leaves are trimmed and treated as waste materials, the trimmed tea leaves are converted into high-value tea leaf fermentation liquor through the method, the utilization rate of tea leaf resources is increased, and greater economic benefits are brought to producers. The pruned tea leaves are converted into the tea leaf fermentation liquor rich in functional components such as tea pigment, theanine and gamma-aminobutyric acid, high practicability is achieved, and the requirements of the market for natural and functional products can be met.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of deep processing of tea tree leaves, and particularly relates to a tea tree leaf fermentation liquor rich in functional components and a preparation method thereof. BACKGROUND

[0002] At present, most of the tea production is still mainly based on "picking tender buds and leaves, discarding small stems and large leaves". Every year, fresh and tender buds and leaves are picked mainly in spring or spring and autumn, and mature leaves and stems (including summer and autumn shoot leaves and current year stems) accounting for 60%-70% of the biomass of tea trees are in a long-term unexploited state due to low picking frequency and difficult processing, especially the huge amount of pruned tea tree leaves are discarded.

[0003] In addition, the difference in metabolic products leads to insufficient umami taste and rough taste of summer and autumn tea, and the quality defects of summer and autumn tea are further highlighted under the traditional tea processing technology, and the market acceptance continues to decline. Influenced by this, the summer and autumn tea gardens in the main tea producing areas in China generally appear the phenomenon of discarding picking, and the pruned tea tree leaves are discarded. The comprehensive utilization rate of tea tree leaves has been less than 10% for a long time. This resource waste has seriously restricted the sustainable development of the tea industry. According to estimates, the fresh weight of leaves and stems produced by pruning in tea gardens can reach 300-500 kg / mu per year, reaching 2574 million tons. The pruned tea tree leaves are rich in tea pigments, theanine, gamma-aminobutyric acid and other active ingredients, and are important biological resources.

[0004] In the prior art, for example, the plant lactobacillus in the patent with publication number CN114933989A can ferment the fermentation medium to obtain a fermentation liquor rich in gamma-aminobutyric acid, and the fermentation liquor can be used to prepare mulberry leaf tea rich in gamma-aminobutyric acid, which provides a new way for the research of mulberry functional food.

[0005] And the patent with publication number CN120131528A is a natural plant fermentation liquor, which is prepared from acanthopanax senticosus, angelica sinensis, rosa davurica, phellinus linteus, tea leaves, water, mixed enzymes and mixed bacterial agents. The mixed enzymes include cellulase, pectinase and tannase, and the mixed bacterial agents include lactobacillus plantarum, schizosaccharomyces and lactobacillus acidophilus. The present application realizes the maximum extraction of the effective active components of natural plants and tea leaves through the synergistic effect of bacteria and enzymes. SUMMARY

[0006] The present application aims to provide a tea tree leaf fermentation liquor rich in tea pigments, theanine, gamma-aminobutyric acid and other functional components and a preparation method thereof. The present application uses a bacteria and enzyme synergistic biological fermentation method to process pruned tea tree leaves to prepare a tea tree leaf fermentation liquor rich in tea pigments, theanine, gamma-aminobutyric acid and other components, which can realize the high-value functional utilization of pruned tea tree leaves and provide a sustainable raw material source for the production of tea tree leaf fermentation liquor functional food.

[0007] The technical solution of this invention: A method for preparing tea leaf fermentation liquid rich in functional components, comprising the following steps: S1: Mix tea leaves with water until evenly mixed to obtain a mixture; S2: Adjust the pH of the mixture to 4-6, add cellulase for enzymatic hydrolysis, and then inactivate the enzyme to obtain tea leaf hydrolysate; S3: Cultivate *Leuciscus roux* ( Zygosaccharomyces rouxii ) and xylospora ( Komagataeibacter xylinus ) bacterial strains; S4: Add sugar to the enzymatic hydrolysate of tea leaves obtained in step S2, and then inoculate it with the microbial strains cultured in step S3 for fermentation. Stir or introduce sterile air during fermentation. After fermentation, perform solid-liquid separation to obtain the tea leaf fermentation broth rich in functional components.

[0008] Preferably, in step S1, the tea leaves are fresh tea leaves or dried tea leaves; The mass ratio of tea tree leaves to water is 1:10-30.

[0009] Furthermore, when the tea leaves are fresh tea leaves, they are pulped to form a slurry. When the tea leaves are dried tea leaves, they are pulverized, and the resulting powder has a mesh size of 20 to 100 mesh.

[0010] Preferably, in step S2, the amount of cellulase added is 40-70 U / mL, based on the volume of the mixed liquid.

[0011] Preferably, in step S2, citric acid solution is used to adjust the pH value of the mixture; The conditions for enzymatic hydrolysis are: stirring at 30-50℃ for 6-12 hours. Enzyme inactivation conditions: Heat to 80-100℃ and hold for 10-30 minutes to inactivate the enzyme.

[0012] The *Lourdescantia rouxae* strain used in this invention ( Zygosaccharomyces rouxii ) is a type of yeast conjugated with Ludwig's yeast ( Zygosaccharomyces rouxii ZUSTZZ-1, accession number CGMCC No. 12131; the described *Xylostella xylostella* ( Komagataeibacter xylinus ) is xylosporinus ( Komagataeibacter xylinus ZUSTKX-1, collection number CGMCC No.31171.

[0013] Preferably, in step S3, the culture time is 24-72 h and the culture temperature is 28-35℃.

[0014] Preferably, in step S4, the sugar is white sugar, brown sugar, honey, maltose, or oligosaccharide; Based on the fresh weight of tea leaf enzymatic hydrolysate, the ratio of added sugar to the mass of tea leaf enzymatic hydrolysate is 0.5-2:1, and the inoculation concentration of *Saccharomyces rouxii* is 1×10⁻⁶. 7 -2×10 7 The inoculation concentration of CFU / mL and *Xylostella xylose* was 8 × 10⁻⁶. 6 -2×10 7 CFU / mL.

[0015] Preferably, in step S4, the fermentation conditions are: fermentation at 28-35℃ for 3-7 days.

[0016] The present invention also provides a tea leaf fermentation liquid rich in functional components prepared by the above preparation method.

[0017] The present invention also provides the application of the above-mentioned tea leaf fermentation liquid rich in functional components in the preparation of functional foods.

[0018] Compared with existing technologies, this invention mixes tea leaves with water and performs enzymatic hydrolysis to obtain tea leaf enzymatic hydrolysate; it then cultivates microbial strains and inoculates the tea leaf enzymatic hydrolysate with microorganisms to obtain a tea leaf fermentation broth rich in theanine, theanine, and γ-aminobutyric acid (GABA). This invention enables the high-value utilization of pruned tea leaves. After the harvesting of tender tea buds and leaves, most tea leaves are pruned and treated as waste. This invention transforms pruned tea leaves into high-value tea leaf fermentation broth, improving the utilization rate of tea leaf resources and bringing greater economic benefits to producers. This invention transforms pruned tea leaves into a tea leaf fermentation broth rich in functional components such as tea pigments, theanine, and GABA, which has strong practicality and can meet the market demand for natural and functional products. Attached Figure Description

[0019] Figure 1 This is a schematic diagram illustrating the preparation process of the tea leaf fermentation liquid rich in functional components according to the present invention. Detailed Implementation

[0020] The present invention will be further described below with reference to embodiments, but these embodiments are not intended to limit the scope of the invention.

[0021] Example 1: Isolation and Identification of Fermentation Strains The *Lourdescantia rouxae* strain used in this invention ( Zygosaccharomyces rouxii Separation occurred in March 2014, and the steps are as follows: 1. Isolation of strains The dragon fruit enzyme stock solution was plated onto PDA agar plates using a serial dilution method to isolate and purify the yeast strain until it was observed to be a pure strain under a microscope. The purified strain was then transferred to an agar slant for storage and named ZUSTZZ-1.

[0022] 2. Morphological identification Colony morphology observation: The purified single colony was inoculated on PDA solid medium and PDA liquid medium, and cultured in a constant temperature incubator at 28°C for 72 h. The colony size, shape, color, whether there was mycelium or island on the liquid surface, whether there was sediment on the bottom layer, etc. were recorded.

[0023] Microscopic observation: A small amount of single colony on PDA solid medium was picked up on a glass slide, diluted with sterile saline, covered with a glass slide, and observed under a microscope at 10x ocular and 100x oil immersion lens. The cell size, shape, and reproduction mode were recorded.

[0024] Scanning electron microscopy: Single colonies were picked up on sterile glass slides, fixed with 2.5% glutaraldehyde solution for 3 h, then washed with 1 / 15 mol / L phosphate buffer for 3 times. Then dehydrated with 50%, 60%, 70%, 80%, 90%, and 100% alcohol solutions for 15 min each time. After treatment, dried in a freeze dryer for 6 h, and used for scanning electron microscopy observation and photography.

[0025] 3. Molecular biology identification According to the operation of Ezup column type yeast genomic DNA extraction kit (SK8257), the universal primer of yeast 26S rDNA gene was used, and the primer sequence was NL-1 (5'-GCATATCAATAAGCGGAGGAAAAG-3') and NL-4 (5'-GGTCCGTGTTTCAAGACGG-3').

[0026] PCR amplification conditions: 94°C, pre-denaturation for 4 min; 94°C, denaturation for 45 s; 55°C, annealing for 45 s; 72°C, extension for 1 min, 30 cycles; 72°C, repair extension for 10 min, 4°C incubation.

[0027] PCR reaction system (25 μL system): Template (genomic DNA 20-50 ng / μL) 0.5 μL; 10×Buffer (containing Mg 2+2.5 μL of dNTPs (2.5 mmol / L each); 1 μL of enzyme; 0.2 μL of forward primer (10 μmol / L); 0.5 μL of reverse primer (10 μmol / L); and finally, add double-distilled water to a final volume of 25 μL. Take 5 μL of the amplification product and perform electrophoresis on a 1% agarose gel at 150 V, 100 mA, for 20 min. Successful amplification will result in a bright band at approximately 600 bp. The sequence obtained from the PCR amplification was determined by Shanghai Sangon Biotech Co., Ltd., and the amplified sequence is shown in SEQ ID NO.1.

[0028] Based on the sequencing results, BLAST software was used to perform a homologous sequence search in the GenBank nucleic acid sequence database (http: / / www.ncbi.nlm.nih.gov / blast) to find and compare homologous or similar nucleotide sequences. Through sequence comparison, strain ZUSTZZ-1 was found to be similar to *Saccharomyces rouxii* (…). Zygosaccharomyces rouxii The sequence is closest to that of *Saccharomyces rouxii*, a result consistent with morphological identification, indicating that the ZUSTZZ-1 strain we isolated is *Saccharomyces rouxii*. Zygosaccharomyces rouxii ).

[0029] The newly screened strain was named *Saccharomyces rouxii*. Zygosaccharomyces rouxii The strain number ZUSTZZ-1 was deposited on February 2, 2016, at the China General Microbiological Culture Collection Center located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 12131. Xylomycin ( Komagataeibacter xylinus Separation occurred in March 2018, and the steps are as follows: 1. Isolation of strains Kombucha culture, provided by the Zhejiang Provincial Key Laboratory of Agricultural Product Chemistry and Bioprocessing Technology, was inoculated into an enrichment medium and cultured at 30°C and 160 r / min for 24 h in a shaker. Using the dilution plating method, the activated bacterial solution was inoculated onto a plate selection medium and cultured at 30°C for 48 h to obtain colonies capable of growth in acetic acid. Single colonies showing good growth on the plates were selected and cultured on acetic acid bacteria solid medium at 30°C for 48 h for strain isolation and purification until the strain was observed to be pure under a microscope. The purified strain was transferred to slant culture for storage and named ZUSTKX-1.

[0030] 2. Morphological characteristics identification Colony morphology observation: The purified single colonies were inoculated into acetic acid bacteria solid culture medium and acetic acid bacteria liquid culture medium, and incubated in a constant temperature incubator at 30℃ for 72 h. The size, shape, color, presence of bacterial spores or islands on the liquid surface, and presence of sediment at the bottom were recorded.

[0031] Microscopic observation: Pick a small amount of single colony from acetic acid bacteria solid culture medium onto a glass slide, dilute with sterile physiological saline, cover with a coverslip, and observe the cell morphology under eyepiece 10× and oil immersion 100× conditions, and record the size, shape and reproduction mode of the bacteria.

[0032] Scanning electron microscopy: Single colonies were picked and placed on sterile glass slides, fixed with 2.5% glutaraldehyde solution for 3 h, and then washed three times with 1 / 15 mol / L phosphate buffer. They were then dehydrated with 50%, 60%, 70%, 80%, 90%, and 100% ethanol solutions, respectively, for 15 min each time. After treatment, they were freeze-dried for 6 h before being observed and photographed under a scanning electron microscope.

[0033] 3. Molecular biological identification The procedure was performed using the Ezup column-based bacterial genomic DNA extraction kit (SK8255), employing universal primers for the bacterial 16S rDNA gene. The primer sequences were: 7F (5´-CAGAGTTTGATCCTGGCT-3´) and 1540R (5´-AGGAGGTGATCCAGCCGCA-3´).

[0034] PCR amplification conditions: 94 ℃, pre-denaturation for 4 min; 94 ℃, denaturation for 45 s; 55 ℃, annealing for 45 s; 72 ℃, extension for 1 min, 30 cycles; 72 ℃, repair extension for 10 min, and incubation at 4 ℃.

[0035] PCR reaction system (25 μL system): Template (genomic DNA 20-50 ng / μL) 0.5 μL; 10× Buffer (containing Mg) 2+ 2.5 μL of dNTPs (2.5 mmol / L each); 1 μL of enzyme; 0.2 μL of forward primer (10 μmol / L); 0.5 μL of reverse primer (10 μmol / L); and finally, add double-distilled water to 25 μL.

[0036] Five μL of the amplification product was subjected to 1% agarose gel electrophoresis at 150 V, 100 mA, for 20 min. Successful amplification was indicated by a bright band at approximately 1500 bp. The sequence obtained from the PCR amplification was determined at Shanghai Sangon Biotech Co., Ltd., and the amplified sequence is shown in SEQ ID NO.2.

[0037] Based on the sequencing results, BLAST software was used to perform a homologous sequence search in the GenBank nucleic acid sequence database (http: / / www.ncbi.nlm.nih.gov / blast) to find and compare homologous or similar nucleotide sequences. Through sequence comparison, strain ZUSTKX-1 was found to be similar to *Xylostella spp.* Komagataeibacter xylinus The sequence is closest to that of *ZUSTKX-1*, a result consistent with morphological identification, indicating that the ZUSTKX-1 strain we isolated is *Z. xylose*. Komagataeibacter xylinus ).

[0038] The newly screened strain was named *Xylosteobacterium xylose* (…). Komagataeibacter xylinus The strain number ZUSTKX-1 was deposited on July 4, 2024, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 31171.

[0039] Example 2 like Figure 1 As shown, a method for preparing tea leaf fermentation broth rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid includes the following steps: S1: Pulp 1 kg of fresh tea leaves into a slurry and mix with water (mass ratio of 1 (dry basis): 20). S2: Adjust the pH of the mixture to 5.0 using citric acid solution, add cellulase and stir under controlled temperature to make the cellulase concentration 40 U / mL, stir at 40℃ for 8 h, then raise the temperature to 90℃ for 20 min to inactivate the enzyme and obtain tea leaf hydrolysate. S3: The *Lourdesca* and *Xylose-containing* strains from Example 1 were cultured for 48 h at a temperature of 30°C. S4: Add granulated sugar (the mass ratio of granulated sugar to tea leaf enzymatic hydrolysate is 1:1) to the tea leaf enzymatic hydrolysate, then inoculate with cultured *Saccharomyces ruuteri* and *Bacillus xylostella*, with an inoculation concentration of 1×10⁻⁶ for both *Saccharomyces ruuteri* and *Bacillus xylostella*. 7 The tea leaf fermentation liquid was obtained by stirring daily for 5 days at a concentration of CFU / mL and a fermentation temperature of 30℃. S5: After fermentation, the tea leaf fermentation liquid is separated into solid and liquid components to obtain tea leaf fermentation liquid rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid.

[0040] Tests showed that the tea pigment content in the tea leaf fermentation liquid was 75 mg / mL, the theanine content was 4.8 mg / mL, and the γ-aminobutyric acid content was 1.5 mg / mL.

[0041] Example 3 A method for preparing tea leaf fermentation broth rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid includes the following steps: S1: Crush 0.5 kg of dried tea leaves to 60 mesh and mix with water (mass ratio of 1 (dry basis): 20); S2: Adjust the pH of the mixture to 4.5 using citric acid solution, add cellulase and stir under controlled temperature to make the cellulase concentration 40 U / mL, stir at 45℃ for 10 h, then raise the temperature to 85℃ for 25 min to inactivate the enzyme, and obtain tea leaf enzymatic hydrolysate. S3: The *Lourdesia rouxae* and *Xylose-bacterium* from Example 1 were cultured for 36 h at a temperature of 28°C. S4: Add brown sugar (brown sugar to tea leaf hydrolysate mass ratio 1.2:1) to the tea leaf hydrolysate, then inoculate with cultured *Saccharomyces ruuteri* and *Bacillus xylostella*. The inoculation concentration of *Saccharomyces ruuteri* is 1.8 × 10⁻⁶. 7 The inoculum concentration of *Bacillus xylostella* was 8 × 10⁻⁶ CFU / mL. 6 The fermentation liquid of tea leaves was obtained by fermenting tea leaves at a concentration of CFU / mL and a fermentation temperature of 28℃ for 7 days with sterile air introduced during fermentation. S5: After fermentation, the tea leaf fermentation liquid is separated into solid and liquid components to obtain tea leaf fermentation liquid rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid.

[0042] Tests showed that the tea pigment content in the tea leaf fermentation liquid was 62 mg / mL, the theanine content was 4.1 mg / mL, and the γ-aminobutyric acid content was 1.8 mg / mL.

[0043] Example 4 A method for preparing tea leaf fermentation broth rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid includes the following steps: S1: Pulp 1.5 kg of fresh tea leaves into a slurry and mix with water (mass ratio of 1 (dry basis): 20). S2: Adjust the pH of the mixture to 6.0 using citric acid solution, add cellulase and stir under controlled temperature to make the cellulase concentration 70 U / mL, stir at 50℃ for 6 h, then raise the temperature to 100℃ for 10 min to inactivate the enzyme and obtain tea leaf enzymatic hydrolysate. S3: Culture the *Lourdesca* and *Xylose-containing* strains from Example 1 for 72 h at a temperature of 30°C. S4: Add honey to the tea leaf enzymatic hydrolysate (honey to tea leaf enzymatic hydrolysate mass ratio is 0.5:1), then inoculate with cultured *Saccharomyces ruvidii* and *Bacillus xylostella*, with an inoculation concentration of 2 × 10⁻⁶ for both *Saccharomyces ruvidii* and *Bacillus xylostella*. 7 The tea leaf fermentation liquid was obtained by fermenting tea leaves at CFU / mL and 30℃ with continuous stirring for 3 days. S5: After fermentation, the tea leaf fermentation liquid is separated into solid and liquid components to obtain tea leaf fermentation liquid rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid.

[0044] Tests showed that the tea pigment content in the tea leaf fermentation liquid was 82 mg / mL, the theanine content was 3.9 mg / mL, and the γ-aminobutyric acid content was 1.2 mg / mL.

[0045] Comparative Example 1 S1: Pulp 1 kg of fresh tea leaves into a slurry and mix with water (mass ratio of 1 (dry basis): 20). S2: After stirring at 40℃ for 8 h, the temperature was raised to 90℃ and heated for 20 min to obtain a tea leaf-water mixture; S3: The *Lourdesca* and *Xylose-containing* strains from Example 1 were cultured for 48 h at a temperature of 30°C. S4: Add white sugar to the tea leaf-water mixture (the mass ratio of white sugar to tea leaf-water mixture is 1:1), then inoculate with cultured *Saccharomyces ruuteri* and *Bacillus xylostella*. The inoculation concentration of both *Saccharomyces ruuteri* and *Bacillus xylostella* is 1×10⁻⁶. 7 The tea leaf fermentation liquid was obtained by stirring daily for 5 days at a concentration of CFU / mL and a fermentation temperature of 30℃. S5: After fermentation, the tea leaf fermentation liquid is separated into solid and liquid components to obtain tea leaf fermentation liquid.

[0046] Tests showed that the tea pigment content in the tea leaf fermentation liquid was 56 mg / mL, the theanine content was 3.6 mg / mL, and the γ-aminobutyric acid content was 0.5 mg / mL.

[0047] Comparative Example 2 S1: Pulp 1 kg of fresh tea leaves into a slurry and mix with water (mass ratio of 1 (dry basis): 20). S2: Adjust the pH of the mixture to 5.0 using citric acid solution, add cellulase and stir under controlled temperature to make the cellulase concentration 40 U / mL, stir at 40℃ for 8 h, then raise the temperature to 90℃ for 20 min to inactivate the enzyme and obtain tea leaf hydrolysate. S3: The *Lourdesian* conjugated yeast from Example 1 was cultured for 48 h at a temperature of 35°C. S4: Add granulated sugar (the mass ratio of granulated sugar to tea leaf hydrolysate is 1:1) to the tea leaf enzymatic hydrolysate, and then inoculate with cultured *Saccharomyces ruvidii* at an inoculation concentration of 1×10⁻⁶. 7 The tea leaf fermentation liquid was obtained by stirring daily for 5 days at a concentration of CFU / mL and a fermentation temperature of 35℃. S5: After fermentation, the tea leaf fermentation liquid is separated into solid and liquid components to obtain tea leaf fermentation liquid.

[0048] Tests showed that the tea pigment content in the tea leaf fermentation liquid was 50 mg / mL, the theanine content was 4.6 mg / mL, and the γ-aminobutyric acid content was 0.4 mg / mL.

[0049] Compared with the comparative examples, the embodiments of the present invention employ a bacterial-enzyme co-fermentation method to obtain a tea leaf fermentation broth rich in functional components such as tea pigments, theanine, and γ-aminobutyric acid. The tea pigment content is 62-82 mg / mL, the theanine content is 3.9-4.8 mg / mL, and the γ-aminobutyric acid content is 1.2-1.8 mg / mL, which is significantly higher than the tea pigment, theanine, and γ-aminobutyric acid content in the tea leaf fermentation broth obtained by the non-enzymatic mixed bacterial fermentation method and the single-strain-enzyme co-fermentation method in the comparative examples.

[0050] In summary, this invention enables the efficient enrichment of functional components such as tea pigments, theanine, and γ-aminobutyric acid (GABA) through synergistic fermentation of bacteria and enzymes, achieving high-value utilization of pruned tea leaves. Pruned tea leaves are typically left directly in the field for composting or discarded without being fully utilized, causing pollution. This invention transforms these low-value, discarded pruned tea leaves into a high-value-added product—a fermented tea leaf liquid rich in functional components such as tea pigments, theanine, and GABA—bringing greater economic benefits to producers. This invention, which converts pruned tea leaves into a fermented tea leaf liquid rich in tea pigments, theanine, and GABA, has strong practicality and can meet the market demand for natural, functional products.

Claims

1. A method for preparing fermented tea leaf broth rich in functional components, characterized in that, Includes the following steps: S1: Mix tea leaves with water until evenly mixed to obtain a mixture; S2: Adjust the pH of the mixture to 4-6, add cellulase for enzymatic hydrolysis, and then inactivate the enzyme to obtain tea leaf hydrolysate; S3: Cultivate *Leuciscus roux* ( Zygosaccharomyces rouxii ) and xylospora ( Komagataeibacter xylinus ) bacterial strains; S4: Add sugar to the enzymatic hydrolysate of tea leaves obtained in step S2, and then inoculate it with the microbial strains cultured in step S3 for fermentation. Stir or introduce sterile air during fermentation. After fermentation, perform solid-liquid separation to obtain the tea leaf fermentation broth rich in functional components.

2. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S1, the tea leaves are either fresh tea leaves or dried tea leaves; The mass ratio of tea leaves to water is 1:10-30.

3. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 2, characterized in that, When the tea leaves are fresh tea leaves, they are pulped to form a slurry. When the tea leaves are dried tea leaves, they are pulverized, and the resulting powder has a mesh size of 20 to 100 mesh.

4. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S2, the amount of cellulase added is 40-70 U / mL, based on the volume of the mixed liquid. The pH of the mixture was adjusted using a citric acid solution. The conditions for enzymatic hydrolysis are: stirring at 30-50℃ for 6-12 hours. Enzyme inactivation conditions: Heat to 80-100℃ and hold for 10-30 minutes to inactivate the enzyme.

5. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S3, the *Leuciscus rouxae* (…) Zygosaccharomyces rouxii ) is a type of yeast conjugated with Ludwig's yeast ( Zygosaccharomyces rouxii ZUSTZZ-1, collection number CGMCC No.12131; The xylosporinus ( Komagataeibacter xylinus ) is xylosporinus ( Komagataeibacter xylinus ZUSTKX-1, collection number CGMCC No.31171.

6. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S3, the incubation time is 24-72 h and the incubation temperature is 28-35℃.

7. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S4, the sugar is white sugar, brown sugar, honey, maltose, or oligosaccharide; Based on the fresh weight of tea leaf enzymatic hydrolysate, the ratio of added sugar to the mass of tea leaf enzymatic hydrolysate is 0.5-2:1, and the inoculation concentration of *Saccharomyces rouxii* is 1×10⁻⁶. 7 -2×10 7 The inoculation concentration of CFU / mL and *Xylostella xylose* was 8 × 10⁻⁶. 6 -2×10 7 CFU / mL.

8. The method for preparing tea leaf fermentation liquid rich in functional components according to claim 1, characterized in that, In step S4, the fermentation conditions are: fermentation at 28-35℃ for 3-7 days.

9. The tea leaf fermentation liquid rich in functional components prepared by the preparation method according to any one of claims 1 to 8.

10. The application of the tea leaf fermentation liquid rich in functional components as described in claim 9 in the preparation of functional foods.

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