Lactiplantibacillus plantarum and use thereof

Abstract

The present invention belongs to the technical field of microorganisms, and specifically relates to a Lactiplantibacillus plantarum, and a composition, culture, fermentation product, food product, or dietary supplement comprising same. The present invention also relates to a method for fermentation using the Lactiplantibacillus plantarum and the composition comprising same. The present invention also relates to use of the Lactiplantibacillus plantarum and the composition comprising same in fermentation and in the preparation of an antioxidant.

Description

A plant lactobacillus and its uses

[0001] Cross-reference to related applications

[0002] This application claims priority to Chinese patent application 202411660138.4, filed on November 20, 2024, the entire contents of which are incorporated herein by reference. Technical Field

[0003] This invention belongs to the field of microbial technology, specifically relating to *Lactobacillus plantarum*, and compositions, cultures, ferments, and food products or dietary supplements containing the same. The invention also relates to methods of fermentation using the *Lactobacillus plantarum* and compositions containing it. Furthermore, the invention relates to the use of the *Lactobacillus plantarum* and compositions containing it in fermentation and the preparation of antioxidants. Background Technology

[0004] Lactiplantibacillus plantarum is a type of lactic acid bacterium widely found in various plants, fermented foods, and different environments. It plays a crucial role in the fermentation process, not only improving the flavor and texture of food but also imparting certain health benefits. Lactiplantibacillus plantarum has significant applications in the food industry, fermentation technology, and biomedicine.

[0005] Kimchi is a traditional pickled vegetable in northern my country, belonging to the category of lactic acid bacteria-fermented foods. Similar to other fermented vegetables, the metabolism of microorganisms not only provides kimchi with rich nutrients but also produces its unique flavor. These flavors mainly include taste substances (organic acids, amino acids, sugars, etc.) and aroma substances (volatile organic acids, alcohols, esters, aromatic compounds, etc.). The fermentation process of kimchi involves the combined action of multiple microorganisms. Research results show that the dominant microorganisms in the fermentation process mainly include *Leuconostoc*, *Weissella*, and *Lactobacillus*. Many of these strains exhibit strong acid and salt tolerance, high temperature tolerance, and strong acid-producing ability, such as *Lactobacillus plantarum*. Among the microorganisms in kimchi, *Leuconostoc* has been widely studied as a starter culture, while *Lactobacillus plantarum* is the dominant microorganism in the later stages of kimchi fermentation, playing a crucial role in the process.

[0006] Current research on *Lactobacillus plantarum* isolated from kimchi mainly focuses on its probiotic properties, with limited research on its application in fermentation. For example, Jeong et al. compared the probiotic properties of *Lactobacillus plantarum* WiKim0112 isolated from kimchi with two other *Lactobacillus plantarum* strains from other sources. The results showed that WiKim0112 exhibited strong acid resistance and resistance to streptomycin and vancomycin, demonstrating potential as a probiotic. Imran Khan studied the functional properties of *Lactobacillus plantarum* DGK-17 isolated from Korean kimchi, investigating its antibacterial and cell invasion protection effects. The results showed that DGK-17 possessed antibacterial activity against various pathogenic microorganisms and could protect HCT 116 colon cells from invasion by target microorganisms. Furthermore, some studies have investigated the effects of *Lactobacillus plantarum* on the fermentation process of kimchi, but these studies involved exogenous *Lactobacillus plantarum* strains rather than its endophytic components. Currently, regarding the application of *Lactobacillus plantarum* isolated from kimchi in other fields, only one study mentions the application of *Lactobacillus plantarum* strain Lb41 isolated from kimchi as a probiotic in cheese. However, this study showed poor fermentation characteristics and failed to improve the fermentation flavor and related physicochemical indicators of the cheese. No research has addressed the application of *Lactobacillus plantarum* from kimchi in other fermented foods.

[0007] Therefore, there is still much room for research on the application of *Lactobacillus plantarum* isolated from kimchi, and the development of a new strain of *Lactobacillus plantarum* suitable for application in the field of fermentation remains to be explored. Summary of the Invention

[0008] The inventors of this application screened a novel strain of *Lactobacillus plantarum* through extensive experiments and, through experimental testing, discovered that it possesses excellent characteristics and significant advantages compared to other *Lactobacillus plantarum* strains (e.g., significant antioxidant capacity, tyrosinase inhibition capacity, and acid-producing capacity, which can significantly improve acid production during fermentation). Thus, the inventors completed this invention.

[0009] Lactobacillus plantarum

[0010] In a first aspect, this application provides a Lactiplantibacillus plantarum, which is deposited at the China General Microbiological Culture Collection Center with accession number CGMCC No. 31269.

[0011] In some embodiments, the *Lactiplantibacillus plantarum* has a 16S rDNA sequence that is 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, 99.95%, or 100% identical to SEQ ID NO:1.

[0012] In some embodiments, the *Lactobacillus plantarum* has the ability to produce more than 1 g / kg (e.g., more than 1.2 g / kg, more than 1.5 g / kg, more than 1.8 g / kg, more than 2.1 g / kg) of total acid under effective culture conditions.

[0013] In some embodiments, the *Lactobacillus plantarum* is able to lower the pH of the fermentation product (e.g., by 0.6 or more).

[0014] In some embodiments, the *Lactobacillus plantarum* can produce a variety of volatile aroma components under effective culture conditions, the flavors being selected from fruity, floral, milky, woody, or any combination thereof.

[0015] In some embodiments, the *Lactobacillus plantarum* is capable of resisting oxidation and / or inhibiting tyrosinase.

[0016] In some embodiments, the *Lactobacillus plantarum* has antioxidant capabilities.

[0017] In some embodiments, the *Lactobacillus plantarum* has a laxative effect that promotes intestinal peristalsis.

[0018] In some embodiments, the *Lactobacillus plantarum* can enhance the nonspecific immune function of the subject. In some embodiments, the *Lactobacillus plantarum* can promote the clearance of carbon particles.

[0019] In some implementations, as mentioned above, *Lactobacillus plantarum* is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 31269.

[0020] In some implementations, the total acid is lactic acid.

[0021] In some embodiments, the fermented product is a fermented food or beverage, such as fermented tea, fermented tea vinegar, fermented fruit juice, or fermented dairy products.

[0022] In some embodiments, the volatile aroma components are selected from acids, esters, alcohols, furans, alkenes, terpenes, or any combination thereof.

[0023] In some embodiments, the volatile aroma component is selected from decanoic acid, 9-decenoic acid, geraniic acid, propionic acid, isoamyl acetate, butyl dodecyl lactone, ethyl octanoate, methyl benzoate, 2,2-dimethyl-5-(1-methyl-1-propenyl)tetrahydrofuran, (E)-β-ocimene, Z-β-ocimene, dipentene, or any combination thereof.

[0024] In some embodiments, the *Lactobacillus plantarum* is capable of scavenging superoxide anions, ABTS radicals, DPPH radicals, hydroxyl radicals, and / or inhibiting superoxide dismutase (SOD).

[0025] In some embodiments, the plant lactobacillus is able to inhibit at least 60% (e.g., at least 60%, at least 65%, at least 70%, at least 75%, at least 80%) of tyrosinase within 3 to 15 minutes (e.g., 3 minutes, 6 minutes, 9 minutes, 12 minutes, 15 minutes).

[0026] Composition

[0027] In a second aspect, this application provides a composition comprising *Lactobacillus plantarum* as described above.

[0028] In some embodiments, the *Lactobacillus plantarum* may be used in combination with one or more other species of microorganisms that can have a beneficial effect on the health of the host to which it is applied.

[0029] Therefore, in some embodiments, the composition further comprises microorganisms selected from bacteria, fungi (e.g., yeast), or any combination thereof, wherein the microorganisms are probiotics.

[0030] In some embodiments, the bacteria are selected from the genera *Lactobacillus*, *C.*, *Bifidobacterium*, *Lactobacillus mucosae*, *Lactobacillus*, *L. assemblica*, *Lactobacillus spp.*, *Streptococcus*, *Lactococcus*, *Propionibacterium*, *Leuconostoc*, *Pediococcus*, *Weizmannella*, *Azoococcus*, *Staphylococcus*, *Bacillus*, *Acetobacter*, *Coprinus*, *Gluconobacterium*, *Gluconobacterium*, or any combination thereof.

[0031] In some embodiments, the bacteria of the genus *Lactobacillus* are selected from: *Lactiplantibacillus plantarum*, *Lactobacillus casei*, *Lactobacillus paracasei*, *Lactobacillus brevis*, *Lactobacillus pentosus*, *Lactobacillus crispatus*, *Lactobacillus rhamnosus*, or any combination thereof.

[0032] In some embodiments, the yeast is selected from Zygosaccharomyces bisporus, Dekkera Bruxellensis, Saccharomyces cerevisiae, Saccharomyces boulardii, Kluyveromyces marxianus, or any combination thereof.

[0033] In some embodiments, the composition as described above comprises *Lactobacillus plantarum* as described above, as well as *Bacillus conjugatus* and / or *Dacobacter brussels*.

[0034] In some embodiments, the *Bacillus conjugatus* is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 31264.

[0035] In some embodiments, the *Dacron brussels yeast* is deposited at the China General Microbiological Culture Collection Center with accession number CGMCC No. 28050.

[0036] In some embodiments, the ratio of *Zygosaccharomyces bisporus*, *Dacronis brusselsii*, and *Lactobacillus plantarum* in the composition is 1-10:1-10:10-20 (e.g., 1:8:16, 1:8:10-20, 1-10:8:16, 1:1-10:16, 1:1-10:10-20, 1-10:8:10-20, 1-10:1-10:16).

[0037] In some embodiments, the ratio of *Bacillus spp.*, *Dacillus brussels*, and *Lactobacillus plantarum* in the composition is 1:8:16.

[0038] In some embodiments, the composition is used for fermentation.

[0039] In some embodiments, the composition further includes additional additives.

[0040] In some embodiments, the additional additive is selected from other nutrients (e.g., carbon sources, nitrogen sources, vitamins, minerals, trace elements, growth factors, or any combination thereof).

[0041] In some embodiments, the carbon source is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof.

[0042] In some embodiments, the carbon source is fruit juice (e.g., grapefruit juice, grape juice, sugarcane juice).

[0043] In some embodiments, the nitrogen source is selected from ammonia, peptone, yeast extract, soybean hydrolysate, milk, or any combination thereof.

[0044] In some embodiments, the additional additives also include tea and / or fruit juice.

[0045] In some embodiments, the tea infusion is selected from black tea infusion, green tea infusion, flower tea infusion, Longjing tea infusion, white tea infusion, eagle tea infusion, oolong tea infusion, barley tea infusion, purple leaf tea infusion, or any combination thereof. In some embodiments, the tea infusion is black tea infusion.

[0046] Culture

[0047] In a third aspect, this application provides a culture comprising *Lactobacillus plantarum* as described above or a composition as described above.

[0048] In some embodiments, the culture also includes additional additives.

[0049] In some embodiments, the additional additive is selected from other nutrients (e.g., carbon sources, nitrogen sources, vitamins, minerals, trace elements, growth factors, or any combination thereof).

[0050] In some embodiments, the carbon source is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof.

[0051] In some embodiments, the carbon source is fruit juice (e.g., grapefruit juice, grape juice, sugarcane juice).

[0052] In some embodiments, the nitrogen source is selected from ammonia, peptone, yeast extract, soybean hydrolysate, milk, or any combination thereof.

[0053] In some embodiments, the additional additives also include tea and / or fruit juice.

[0054] In some embodiments, the tea infusion is selected from black tea infusion, green tea infusion, flower tea infusion, Longjing tea infusion, white tea infusion, eagle tea infusion, oolong tea infusion, barley tea infusion, purple leaf tea infusion, or any combination thereof.

[0055] In some implementations, the tea infusion is black tea infusion.

[0056] In some embodiments, the culture is a solid, liquid, or semi-solid.

[0057] In some embodiments, the culture further comprises a cell-free culture filtrate of the *Lactobacillus plantarum*.

[0058] In some embodiments, the culture also contains a derivative of Lactobacillus plantarum.

[0059] In some embodiments, the derivative is selected from metabolites, supernatants of *Lactobacillus plantarum*, lysates of *Lactobacillus plantarum*, or any combination thereof.

[0060] In some embodiments, the supernatant of *Lactobacillus plantarum* is the supernatant obtained after centrifuging a culture of *Lactobacillus plantarum*. In some embodiments, the lysate of *Lactobacillus plantarum* is the supernatant obtained after lysing the bacterial cells of *Lactobacillus plantarum*.

[0061] Food products or dietary supplements

[0062] In a fourth aspect, this application provides a food product or dietary supplement comprising *Lactobacillus plantarum* as described above, or a composition or culture as described above.

[0063] In some implementations, the food product or dietary supplement is a beverage.

[0064] In some embodiments, the food product or dietary supplement is selected from tea drinks, solid beverages, or fruit juice drinks.

[0065] In some embodiments, the food product or dietary supplement comprises dairy products (e.g., yogurt, flavored fermented milk, lactic acid bacteria beverages, cheese).

[0066] In some implementations, the food product or dietary supplement may also include one or any combination of the following substances: probiotics (e.g., probiotics), dietary fiber, prebiotics, proteins (e.g., enzymes), carbohydrates, lipids (e.g., fats), vitamins, minerals, plant ingredients (e.g., plant extracts), amino acids, immunomodulators, and milk substitutes.

[0067] In a fifth aspect, this application provides a fermentation method comprising: inoculating a tea infusion with *Lactobacillus plantarum* as described above or a composition as described above. In some embodiments, *Lactobacillus plantarum* is cultured in the inoculated tea infusion. In some embodiments, the tea infusion refers to tea water, filtrate, or extract after brewing tea leaves.

[0068] In a sixth aspect, this application provides a fermentation product obtained by fermentation via *Lactobacillus plantarum* as described above, or by fermentation via the composition or culture described above, or by fermentation method described above.

[0069] In some embodiments, the fermented product is fermented tea, fermented tea vinegar, fermented fruit juice, fermented dairy products, or any combination thereof.

[0070] In some embodiments, the fermentation product also includes additional additives.

[0071] In some embodiments, the additional additives are selected from sweeteners, colorings, preservatives, antioxidants, acidity regulators, nutritional fortifiers, or any combination thereof.

[0072] In some embodiments, the fermented tea is prepared by fermentation using Lactobacillus plantarum as described above, or it is obtained by the fermentation method described above.

[0073] In some embodiments, the tea is selected from black tea, green tea, flower tea, Longjing tea, white tea, eagle tea, oolong tea, barley tea, or any combination thereof.

[0074] In some implementations, the tea is black tea.

[0075] In some embodiments, the tea content is 0.2%-1.6% (m / v) (e.g., 0.2%, 0.4%, 0.8%, 1.2%, 1.6%).

[0076] In some embodiments, the tea content is 0.8% (m / v).

[0077] In some implementations, the total sugar content (Brix%) of fermentation is 2%-8% (m / v) (e.g., 2%, 4%, 6%, 8%).

[0078] In some implementations, the total sugar content (Brix%) of fermentation is 4% (m / v).

[0079] In some implementations, the carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof.

[0080] In some implementations, the carbon source for fermentation is selected from sucrose, glucose, grapefruit juice, or any combination thereof.

[0081] In some implementations, the carbon source for fermentation is sucrose, glucose, and grapefruit juice in a ratio of 4:2:3.

[0082] In some implementations, the carbon source for fermentation is sucrose, glucose, and grapefruit juice, with the addition amounts being 2% sucrose, 1% glucose, and 1.5% grapefruit juice.

[0083] In some implementations, the fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days).

[0084] In some implementations, the fermentation time is 3 days.

[0085] In some embodiments, the fermented tea has a lemony aroma.

[0086] In some embodiments, the fermented tea vinegar is prepared by fermentation of yeast containing *Gnaphalium affine*, *Dacron brückensis*, and *Lactobacillus plantarum* as described above, or it is obtained by the fermentation method described above.

[0087] In some embodiments, the *Bacillus conjugatus* is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 31264.

[0088] In some embodiments, the *Dacron brussels yeast* is deposited at the China General Microbiological Culture Collection Center with accession number CGMCC No. 28050.

[0089] In some embodiments, the ratio of *Gnaphalium affine*, *Dacronis brusselsii*, and *Lactobacillus plantarum* is 1-10:1-10:10-20 (e.g., 1:8:16, 1:8:10-20, 1-10:8:16, 1:1-10:16, 1:1-10:10-20, 1-10:8:10-20, 1-10:1-10:16).

[0090] In some embodiments, the ratio of the *Dictyophora indicum*, *Dictyophora brusselii*, and *Lactobacillus plantarum* is 1:8:16.

[0091] In some embodiments, the tea vinegar has a pH value of less than 4 (e.g., pH value less than 3.6, pH value less than 3.2).

[0092] In some embodiments, the tea in the tea vinegar is selected from black tea, green tea, flower tea, Longjing tea, white tea, eagle tea, oolong tea, barley tea, or any combination thereof.

[0093] In some embodiments, the tea in the tea vinegar is black tea.

[0094] In some embodiments, the tea content is 0.2%-1.6% (m / v) (e.g., 0.2%, 0.4%, 0.8%, 1.2%, 1.6%).

[0095] In some embodiments, the tea content is 0.8% (m / v).

[0096] In some implementations, the total sugar content (Brix%) of fermentation is 2%-8% (m / v) (e.g., 2%, 4%, 6%, 8%).

[0097] In some implementations, the total sugar content (Brix%) of fermentation is 4% (m / v).

[0098] In some implementations, the carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof.

[0099] In some implementations, the carbon source for fermentation is selected from sucrose, glucose, grapefruit juice, or any combination thereof.

[0100] In some implementations, the ratio of sucrose, glucose, and grapefruit juice as the carbon source for fermentation is 4:2:3.

[0101] In some implementations, the amount of sucrose, glucose, and grapefruit juice added as carbon sources for fermentation is 2% sucrose, 1% glucose, and 1.5% grapefruit juice.

[0102] In some implementations, the fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days).

[0103] In some implementations, the fermentation time is 3 days.

[0104] In some embodiments, the fermented fruit juice is prepared by fermentation containing *Lactobacillus plantarum* as described above, or it is obtained by the fermentation method described above.

[0105] In some implementations, the carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof.

[0106] In some implementations, the carbon source for fermentation is white grape juice.

[0107] In some implementations, the carbon source for fermentation is 5%-10% white grape juice (e.g., 8.1%).

[0108] In some implementations, the fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days).

[0109] In some implementations, the fermentation time is 3 days.

[0110] In some implementations, the fermentation temperature is 23–32°C (e.g., 24°C, 26°C, 28°C, 30°C).

[0111] In a seventh aspect, this application provides the use of *Lactobacillus plantarum* as described above, or the composition or culture described above, in fermentation.

[0112] In some embodiments, the fermentation is used to prepare fermented tea, fermented tea vinegar, fermented fruit juice, fermented dairy products, or any combination thereof.

[0113] In an eighth aspect, this application provides the use of *Lactobacillus plantarum* as described above, or the composition or culture described above, in the preparation of antioxidants.

[0114] In some embodiments, the antioxidant is able to inhibit tyrosinase.

[0115] In some embodiments, the antioxidant is capable of scavenging superoxide anions, scavenging ABTS radicals, scavenging DPPH radicals, scavenging hydroxyl radicals, and / or inhibiting superoxide dismutase (SOD).

[0116] In a ninth aspect, this application provides the use of *Lactobacillus plantarum* as described above, or the composition or culture described above, in the preparation of food products or dietary supplements.

[0117] In some implementations, the food product or dietary supplement is a beverage.

[0118] In some embodiments, the food product or dietary supplement is selected from tea drinks, solid beverages, or fruit juice drinks.

[0119] In some embodiments, the food product or dietary supplement comprises dairy products (e.g., yogurt, flavored fermented milk, lactic acid bacteria beverages, cheese).

[0120] In a tenth aspect, this application provides the use of *Lactobacillus plantarum* as described above, or the composition or culture as described above, in the preparation of a medicament for the prevention and / or treatment of constipation in a subject, or for enhancing the immune function (e.g., nonspecific immunity) of a subject.

[0121] In some embodiments, the drug has a laxative effect. In some embodiments, the drug can promote intestinal peristalsis. In some embodiments, the drug can promote the clearance of carbon particles from the subject.

[0122] Terminology Definition

[0123] As used herein, the term "fermentation" is a biochemical reaction process involving the conversion of organic matter into energy, gases, and other bioactive substances (e.g., including but not limited to organic acids, esters, and aroma compounds) by microorganisms (e.g., bacteria and / or yeast) under anaerobic or hypoxic conditions. This process can be used to prepare the microbial cells themselves, or direct or secondary metabolites. Fermentation can occur spontaneously or under controlled industrial production conditions. In some embodiments, the fermentation is used to prepare fermented tea, fermented tea vinegar, and fermented fruit juice. The term "fermentation product" refers to the substance obtained through fermentation.

[0124] As used in this article, the term "total acidity" refers to the total amount of all acidic components in a food, including those that have dissociated into H+. + Total acid is the concentration of acids (free state) and undissociated acids (bound state, acid salts). Total acid is present in fruit and vegetable products, beverages, dairy products, wines, grain products, and condiments. The determination of total acid is one of the key indicators for evaluating fermentation efficiency and product quality. It reflects the total content of organic weak acids in food, such as lactic acid, malic acid, citric acid, and acetic acid. In some implementations, total acid refers to the content of lactic acid.

[0125] As used herein, the term "tea infusion" is a liquid obtained by mixing tea leaves with hot water in a specific ratio, extracting the mixture at a specific temperature for a specific time, and then filtering to remove tea residue. Tea infusion contains not only the water-soluble components of the tea leaves (e.g., tea polyphenols, caffeine, and amino acids) but also the aroma and flavor of the tea. In some embodiments, the tea leaves are black tea. In some embodiments, the tea infusion is used to inoculate microorganisms (e.g., bacteria and / or yeast) for fermentation.

[0126] As used herein, the term "tea vinegar" refers to a beverage-type vinegar product made primarily from tea leaves through methods such as extraction or bio-fermentation. In some embodiments, bacteria and yeast are used for bio-fermentation.

[0127] As used herein, the term "identity" refers to the sequence matching between two polypeptides or two nucleic acids. Two compared sequences are identical at a position when the same base or amino acid monomeric subunit occupies the same location (e.g., a position in each of two DNA molecules is occupied by adenine, or a position in each of two polypeptides is occupied by lysine). The "percentage identity" between two sequences is a function of the number of matching positions shared by the two sequences divided by the number of positions compared × 100. For example, if six out of ten positions in two sequences match, then the two sequences have 60% identity. For example, the DNA sequences CTGACT and CAGGTT share 50% identity (three out of six positions match). Typically, two sequences are compared to produce the maximum identity. Such comparisons can be made using methods readily available, for example, computer programs such as the Align program (DNAstar, Inc.) Needleman et al. (1970) J. Mol. Biol. 48: 443-453. The percentage identity between two amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl Biosci., 4:11-17 (1988)) integrated into the ALIGN program (version 2.0), which uses a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Alternatively, the percentage identity between two amino acid sequences can be determined using the Needleman and Wunsch algorithm (J MoI Biol. 48:444-453 (1970)) in the GAP program integrated into the GCG software package (available at www.gcg.com), which uses a Blossum 62 matrix or a PAM250 matrix, along with gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0128] Beneficial effects of the invention

[0129] The *Lactobacillus plantarum* strain of this application has excellent characteristics and, compared with other *Lactobacillus plantarum* strains, has at least the following advantages: (1) it has significant antioxidant capacity and does not cause toxic side effects to the subjects; (2) it has tyrosinase inhibitory capacity and can significantly inhibit it in a short time; (3) it has acid production capacity and can significantly increase acid production during fermentation; (4) it has a laxative effect that promotes intestinal peristalsis; and (5) it can enhance the non-specific immune function of the subjects and promote the clearance of carbon particles.

[0130] Furthermore, the inventors of this application have demonstrated through extensive experiments that this *Lactobacillus plantarum* can grow efficiently and produce high levels of acid (e.g., at least 1.2 g / kg) under various fermentation conditions. It exhibits efficient growth and high acid production in the fermentation of tea, tea vinegar, and fruit juice, and also improves the flavor and sensory quality of the fermented products.

[0131] The embodiments of the present invention will now be described in detail with reference to the accompanying drawings and examples. However, those skilled in the art will understand that the following drawings and examples are for illustrative purposes only and are not intended to limit the scope of the invention. Various objects and advantages of the present invention will become apparent to those skilled in the art from the following detailed description of the drawings and preferred embodiments. Attached Figure Description

[0132] Figure 1 shows the sensory evaluation results of the four lactic acid bacteria strains.

[0133] Figure 2 shows the pH changes in the fermentation broth when four different lactic acid bacteria were mixed with yeast during the fermentation of fermented tea vinegar. From left to right, the five experimental groups contain the following lactic acid bacteria: no lactic acid bacteria added, *Lactobacillus plantarum* XS001, *Lactobacillus plantarum* L-M23-6, *Lactococcus lactis* FZ022, and *Lactobacillus plantarum* L-Y7-6.

[0134] Figure 3 shows the OD values ​​of the fermentation broth when four different lactic acid bacteria were mixed with yeast during the fermentation of fermented tea vinegar. 600 Changes. From left to right, the five experimental groups contained the following lactic acid bacteria: no lactic acid bacteria added, *Lactobacillus plantarum* XS001, *Lactobacillus plantarum* L-M23-6, *Lactococcus lactis* FZ022, and *Lactobacillus plantarum* L-Y7-6.

[0135] Figure 4 shows the changes in total acid content (TA) of the fermentation broth when four different lactic acid bacteria were mixed with yeast during the fermentation of fermented tea vinegar. From left to right, the five experimental groups contained the following lactic acid bacteria: no lactic acid bacteria added, *Lactobacillus plantarum* XS001, *Lactobacillus plantarum* L-M23-6, *Lactococcus lactis* FZ022, and *Lactobacillus plantarum* L-Y7-6.

[0136] Figure 5 shows the sensory evaluation results of fermented tea vinegar after 4 days of fermentation.

[0137] Figure 6 shows the changes in sugar content of fermentation broth when Lactobacillus plantarum L-Y7-6 ferments fruit juice with different carbon sources.

[0138] Figure 7 shows the changes in total acid content of fermentation broth when Lactobacillus plantarum L-Y7-6 ferments fruit juice with different carbon sources.

[0139] Figure 8 shows the OD values ​​of the fermentation broth from Lactobacillus plantarum L-Y7-6 fermenting fruit juices with different carbon sources. 600 Changes.

[0140] Figure 9 shows the sensory evaluation results of different fermented fruit juices by Lactobacillus plantarum L-Y7-6.

[0141] Figure 10 shows the volatile aroma components of fermented tea as determined by GC-MS.

[0142] Sequence information

[0143] Information on some of the sequences involved in this invention is provided in Table 1 below.

[0144] Table 1: Sequence Description

[0145] Instructions on the Preservation of Biological Materials

[0146] Lactiplantibacillus plantarum L-Y7-6 has been deposited at the China General Microbiological Culture Collection Center (CGMCC) located on Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 31269 and deposit date of July 11, 2024. Detailed Implementation

[0147] The invention will now be described with reference to the following embodiments, which are intended to illustrate the invention (and not limit it).

[0148] Unless otherwise specified, the experiments and methods described in the embodiments are performed in accordance with conventional methods well known in the art and described in various references.

[0149] Furthermore, unless specific conditions are specified in the examples, conventional conditions or conditions recommended by the manufacturer should be followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products. Those skilled in the art will understand that the examples are described by way of illustration and are not intended to limit the scope of protection claimed by the invention. All disclosures and other references mentioned herein are incorporated herein by reference in their entirety.

[0150] Example 1. Obtaining strains and applying four types of lactic acid bacteria in fermented tea.

[0151] (1) Isolation, purification and preliminary identification of lactic acid bacteria

[0152] The collected Yanji kimchi juice samples were serially diluted with sterile physiological saline and inoculated into solid MRS medium containing 1.5% calcium carbonate. After anaerobic incubation at 37°C for 48-72 hours, colonies with calcium rings, round shape, neat edges, smooth and moist surface, and opaque appearance were selected. After streak isolation and purification 2-3 times, single colonies were selected, and Gram staining and microscopic examination were performed to confirm pure culture, Gram-positive non-spore-forming bacteria. Single colonies were then inoculated into liquid medium and incubated in an anaerobic workstation at 37°C for 24-48 hours to obtain pure cultures, which were then frozen in 25% glycerol at -80°C.

[0153] (2) Identification of lactic acid bacteria species

[0154] Lactic acid bacteria DNA was extracted using a kit method. Using this DNA as a template, PCR amplification of the 16S rDNA of the candidate *Lactiplantibacillus plantarum* was performed using universal 16S rDNA primers (forward primer 27F: SEQ ID NO:2; reverse primer 1495R: SEQ ID NO:3). The product was sent to Sangon Biotech for sequencing, and the result is shown in SEQ ID NO:1. The obtained sequence was submitted to GenBank for BLAST comparison analysis. The BLAST comparison results showed that the sequence of L-Y7-6 had more than 99% homology with the 16S rDNA sequence of *Lactiplantibacillus plantarum*.

[0155] Therefore, the applicant obtained several novel strains of *Lactobacillus plantarum* and deposited them into a library, naming them L-M23-6, XS001, XS002, XS004, and L-Y7-6, respectively. Furthermore, these *Lactobacillus plantarum* strains were tested as described in the examples below, and *Lactobacillus plantarum* strain L-Y7-6 was found to have outstanding activity among the numerous strains. Consequently, the applicant deposited L-Y7-6 on July 11, 2024.

[0156] Application of 4 types of lactic acid bacteria in fermented tea

[0157] 1.1 Experimental Materials

[0158] Experimental strains: see Table 1-1.

[0159] Table 1-1: Experimental bacterial strains

[0160] Laboratory supplies: pH meter (Mettler, Sz-standard); saccharimeter (Aitol, RL-1); potentiometric titrator (Mettler, TITRATOR) T50); Full-wavelength microplate reader (Epoch2, equipment number 70000817); Ultraviolet spectrophotometer (Shimadzu, Japan, 70001618); High-temperature steam sterilizer (Puhexi Co., Ltd., MLS-3781L-PC); Incubator (Yonglian Biotechnology, UL3L-12); Analytical balance (Mettler, SP029); Clean bench (equipment number YST-YJS-0025); Centrifuge (5810R / Eppendorf, equipment number 70000218); Sucrose (230630, Guangxi Chongzuo Dongya Sugar Industry Co., Ltd.); Glucose (Weifang Shengtai); Grapefruit juice (230620-#4, Nongfu Spring); MRS culture medium (Qingdao Haibo, batch number 20210603).

[0161] 1.2 Experimental Scheme

[0162] 1) Activation of bacterial strains: Take L-Y7-6, XS001, L-M23-6 and Lactococcus lactis FZ022 preserved in glycerol tubes and inoculate them into MRS medium at an inoculation rate of 1.0% and incubate at 37°C for 24 hours.

[0163] 2) Preparation of fermentation starter culture: In a clean bench, take 20 mL of the bacterial culture prepared in step 1) and place it in a 50 mL centrifuge tube. Centrifuge at 8000 rpm for 10 min. After centrifugation, discard the supernatant. Add approximately 25 mL of sterilized tea broth to the centrifuge tube to resuspend the bacterial culture. Centrifuge again at 8000 rpm for 10 min. Finally, discard the supernatant and add another 20 mL of sterilized tea broth to prepare the seed culture. The sterilized tea broth is obtained through extraction in step 3) and sterilization in step 5).

[0164] 3) Tea extraction: Weigh a certain amount of black tea B into a preheated ceramic jar, add the corresponding amount of 85℃ hot water at a tea-to-water ratio of 1:30, heat in a water bath at 85℃ for 15.0 min, stirring once every 5 min, and after heating, quickly filter out the tea leaves and tea residue with a filter screen, and put the tea in an ice water bath to quickly cool down and lock in the aroma.

[0165] 4) Preparation of Tea Sugar Water: After the tea infusion has cooled to room temperature, prepare the tea sugar water. Based on a total sugar content of approximately 4.1% (Brix%) (2% sucrose + 1% glucose + 1.5% concentrated grapefruit juice), weigh out a certain amount of carbon source into a stainless steel beaker. Add concentrated extract of black tea B (20%) according to a tea content of 0.8%, and bring to a certain mass, then stir well. Next, weigh 400.0g of the tea sugar water into an Erlenmeyer flask and seal it with double-layered aluminum foil and sealing film.

[0166] 5) Sterilization: The prepared tea sugar water is pasteurized in an autoclave (70℃ for 15 min).

[0167] 6) Inoculation: After the tea and sugar water is sterilized, cool it to room temperature. Inoculate each type of lactic acid bacteria in a laminar flow hood at a 1% inoculation concentration of the seed liquid obtained in step 2), and then mix it in an Erlenmeyer flask.

[0168] 7) Fermentation: The conical flasks were placed in a shaker at 37℃ and 120 rpm for cultivation. The specific fermentation process is shown in Table 1-2.

[0169] 8) Sampling: Determine the sugar content (Brix%), pH, and total acid of the initial tea sugar solution (0h); take 50mL samples at 24h, 48h, and 72h of fermentation respectively, and determine the OD. 600 The sugar content (Brix%), pH, and total acid were measured, and sensory evaluations were conducted to explore changes in fermentation flavor.

[0170] Table 1-2: Fermentation Process Parameters

[0171] 1.3 Experimental Results

[0172] The OD values ​​of the initial tea sugar solution (0h) and samples fermented for 24h, 48h, and 72h were measured. 600 The results for pH, sugar content (Brix%), and total acid are shown in Tables 1-3 to 1-6 below.

[0173] Table 1-3: Oddi concentration of fermented tea 600 change

[0174] Table 1-4: pH Changes in Fermented Tea

[0175] Table 1-5: Brix Variations in Fermented Tea

[0176] Table 1-6: Changes in total acidity in fermented tea (g / kg) Note: "Blank cells" indicate that no measurement is required based on sensory results.

[0177] Experimental results

[0178] 1. Fermentation results

[0179] In the fermented tea system, L-Y7-6 showed the fastest growth compared to other strains, with an OD of [missing value] after 72 hours of fermentation. 600The pH reached 0.483; it had the strongest acid-producing capacity, the lowest pH at 72 hours of fermentation (3.33), and the highest total acidity (2.34), exhibiting the highest batch-to-batch stability. Flavor evaluation of fermented tea showed that strain L-Y7-6 fermentation broth was rich in acidity and fruity aroma, with a strong lemon-black tea flavor, and demonstrated the strongest flavor stability across multiple fermentation batches.

[0180] 2. Sensory evaluation results

[0181] Sensory evaluation involved 15 participants. Each participant drank all five test samples in one round, with a 120-second rest between each sample. Samples were filtered with water and Sudan biscuits between rounds. Flavor evaluation focused on four aspects: aroma, taste, mouthfeel, and defects, with a final overall preference rating. The overall preference rating used a 9-point scale: 1 point - extremely dislike; 5 points - neither dislike nor dislike; 9 points - extremely like. Generally, 5.4 points was considered acceptable, and 7.2 points was excellent. The overall scoring results are shown in Figure 1. The L-Y7-6 fermentation broth was rich in acidity and fruity aroma, with a strong lemon-black tea flavor. The flavor of the 3-day fermented broth was better than that of the 2-day broth. At 3 days of fermentation, the sensory evaluation results of *Lactobacillus plantarum* L-Y7-6 from this application were significantly better than other strains, and it exhibited the strongest flavor stability across multiple batches of fermentation.

[0182] Example 2. Application of Lactobacillus plantarum L-Y7-6 in fermented tea

[0183] 2.1 Experimental Materials

[0184] Experimental strain: Lactobacillus plantarum L-Y7-6.

[0185] Laboratory supplies: pH meter (Mettler, Sz-standard); saccharimeter (Aitol, RL-1); potentiometric titrator (Mettler, TITRATOR) T50); Full-wavelength microplate reader (Epoch2, equipment number 70000817); Ultraviolet spectrophotometer (Shimadzu, Japan, 70001618); High-temperature steam sterilizer (Puhexi Co., Ltd., MLS-3781L-PC); Incubator (Yonglian Biotechnology, UL3L-12); Analytical balance (Mettler, SP029); Clean bench (equipment number YST-YJS-0025); Centrifuge (5810R / Eppendorf, equipment number 70000218); Sucrose (230630, Guangxi Chongzuo Dongya Sugar Industry Co., Ltd.); Glucose (Weifang Shengtai); Grapefruit juice (230620-#4, Nongfu Spring); MRS medium (Qingdao Haibo, batch number 20210603).

[0186] 2.2 Experimental Scheme

[0187] 1) Activation of bacterial strain: Take L-Y7-6 of Lactobacillus plantarum preserved in glycerol tubes, inoculate it into MRS medium at an inoculation rate of 1.0%, and incubate at 37°C for 24 hours.

[0188] 2) Preparation of fermentation starter culture: In a clean bench, take 20 mL of the bacterial culture prepared in step 1) and place it in a 50 mL centrifuge tube. Centrifuge at 8000 rpm for 10 min. After centrifugation, discard the supernatant. Add approximately 25 mL of sterilized tea broth to the centrifuge tube to resuspend the bacterial culture. Centrifuge again at 8000 rpm for 10 min. Finally, discard the supernatant and add another 20 mL of sterilized tea broth to prepare the seed culture. The sterilized tea broth is obtained through extraction in step 3) and sterilization in step 5).

[0189] 3) Tea extraction: Weigh a certain amount of black tea B into a preheated ceramic jar, add the corresponding amount of 85℃ hot water at a tea-to-water ratio of 1:30, heat in a water bath at 85℃ for 15.0 min, stirring once every 5 min, and after heating, quickly filter out the tea leaves and tea residue with a filter screen, and put the tea in an ice water bath to quickly cool down and lock in the aroma.

[0190] 4) Preparation of Tea Sugar Water: After the tea infusion has cooled to room temperature, prepare the tea sugar water. Based on a total sugar content of 4.1% (Brix%) (2% sucrose + 1% glucose + 1.5% concentrated grapefruit juice), weigh out a certain amount of carbon source into a stainless steel beaker. Add concentrated extract of black tea B, based on a tea content of 0.8%, and bring to a certain mass. Then stir well. Next, weigh 400.0g of the tea sugar water into an Erlenmeyer flask and seal it with double-layered aluminum foil and sealing film.

[0191] 5) Sterilization: The prepared tea sugar water is pasteurized in an autoclave (70℃ for 15 min).

[0192] 6) Inoculation: After the tea sugar water is sterilized, cool it to room temperature, inoculate the seed liquid obtained in step 2) at a certain inoculation concentration in the laminar flow hood, and then mix it in an Erlenmeyer flask.

[0193] 7) Fermentation: The conical flasks were placed in a shaker at 37℃ and 120rpm for cultivation. The specific fermentation process is shown in Table 2-1.

[0194] 8) Sampling: The initial tea sugar water (0h) needs to be tested for sugar content (Brix%), pH, total acid and tea polyphenols; 50mL samples are taken at 48h and 72h of fermentation respectively, and the sugar content (Brix%), pH, total acid and tea polyphenol content (72h sample) are tested. At the same time, sensory evaluation is carried out to study the changes in fermentation flavor.

[0195] Table 2-1: Fermentation Process Parameters

[0196] 2.3 Experimental Results

[0197] 1. Fermentation results

[0198] Table 2-2 Indicators of fermented tea containing Lactobacillus plantarum L-Y7-6

[0199] Note: Total acid content is calculated as lactic acid (same below).

[0200] As the fermentation time increases, the acidity of the fermented tea increases slightly.

[0201] 2. Sensory evaluation results

[0202] Sensory evaluation involved 15 participants. Each participant drank all five test samples in one round, with a 120-second rest between each sample. Samples were filtered with water and Sudan biscuits between rounds. Flavor evaluation focused on four aspects: aroma, taste, mouthfeel, and defects, with a final overall preference rating. The overall preference rating used a 9-point scale: 1 point - extremely dislike; 5 points - neither dislike nor dislike; 9 points - extremely like. A score of 5.4 was generally considered acceptable, and 7.2 points - excellent. Overall scores showed that with prolonged fermentation, the acidity of the fermented tea increased slightly, and the flavor gradually became richer, exhibiting floral, fruity, milky, and lemon-black tea notes. The fermented liquid after 3 days showed better flavor than that after 2 days and 1 day, with sensory evaluation scores of 6.0 (3 days), 5.38 (2 days), and 4.0 (1 day), respectively.

[0203] 3. Results of volatile component analysis

[0204] The volatile aroma components were determined by GC-MS (Figure 10), showing that Lactobacillus plantarum L-Y7-6 can metabolize and produce 13 acids, 15 alcohols, 8 esters, 3 phenols, 5 alkenes, and 2 terpenes, totaling 53 volatile components, of which as many as 12 are related to fruity aroma.

[0205] Example 3.4 Application of lactic acid bacteria in fermented tea vinegar

[0206] 3.1 Experimental Materials

[0207] Experimental strains: see Table 3-1.

[0208] Table 3-1: Experimental bacterial strains

[0209] Among them, *Gnaphalium zygosacchari* YF9-1 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 31264; *Dacron brüsseldorf* Y5-6 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 28050.

[0210] Laboratory supplies: pH meter (Mettler, Sz-standard); saccharimeter (Aito, RL-1); potentiometric titrator (Mettler, TITRATOR T50); full-wavelength microplate reader (Epoch2, equipment number 70000817); UV spectrophotometer (Shimadzu, Japan, 70001618); high-temperature steam sterilizer (Puwashi Co., Ltd., MLS-3781L-PC); incubator (Yonglian Bio, UL3L-12); analytical balance (Mettler, SP029); clean bench (equipment number YST-YJS-0025); centrifuge (5810R / Eppendorf, equipment number 70000218).

[0211] Black tea B (230419 / 230602), sucrose (230630, Guangxi Chongzuo Dongya Sugar Industry Co., Ltd.); glucose (Weifang Shengtai); grapefruit juice (230620-#4, Nongfu Spring); YPD medium (Qingdao Haibo, batch number 20210603); MRS medium (Qingdao Haibo, batch number 20210603).

[0212] 3.2 Experimental Scheme

[0213] (1) Activation of strains: 1.0 mL (2.0%) of yeast strain YF9-1 and yeast strain Y5-6 preserved in glycerol tubes were inoculated into 50 mL of YPD medium and then cultured at 28℃ and 180 rpm for 2 days. 1.0% of three types of Lactobacillus plantarum L-Y7-6, XS001, L-M23-6 and Lactococcus lactis FZ022 preserved in glycerol tubes were inoculated into MRS medium and cultured at 37℃ for 24 h.

[0214] (2) Preparation of fermentation strains: In a clean bench, 40.0 mL of the two yeast cultures and four lactic acid bacteria cultures of *Lactobacillus plantarum* as shown in step (1) were respectively placed into 50.0 mL centrifuge tubes. The tubes were then centrifuged at 6000 rpm for 15 min. After centrifugation, the supernatant was discarded, and about 45.0 mL of sterilized tea broth was added to the centrifuge tubes to resuspend the bacterial culture. The tubes were then centrifuged again at 6000 rpm for 15.0 min. Finally, the supernatant was discarded, and another 40.0 mL of sterilized tea broth was added to prepare the seed culture. The sterilized tea broth was obtained through extraction in step (3) and sterilization in step (5).

[0215] (3) Tea extraction: Weigh a certain amount of black tea B into a preheated ceramic jar, add the corresponding amount of hot water at 85℃ according to a tea-to-water ratio of 1:30, heat in a water bath at 85℃ for 15.0 min, stir once every 5 min, and after heating, quickly filter out the tea leaves and tea residue with a filter screen, and put the tea into an ice water bath to quickly cool down and lock in the aroma.

[0216] (4) Preparation of tea syrup: After the tea soup has cooled to room temperature, prepare the tea syrup. Weigh out a certain amount of carbon source (2% sucrose + 1% glucose + 1.5% concentrated grapefruit juice) into a stainless steel beaker according to a total sugar content of 4.0% (Brix, %). Add concentrated extract of black tea B according to a tea content of 0.8%, bring to a certain mass, and then stir well. Next, weigh 400.0g of the tea syrup into an Erlenmeyer flask and seal it with double-layered aluminum foil and sealing film.

[0217] (5) Sterilization: The prepared tea sugar water is pasteurized in an autoclave (70℃ for 15 min).

[0218] (6) Inoculation: After the tea sugar water is sterilized, it is cooled to room temperature. The seed liquid obtained in step (2) is inoculated in a clean bench according to different inoculation combinations and ratios (Y5-6:YF9-1: Lactic acid bacteria (L-Y7-6, XS001, L-M23-6, FZ002) = 8:1:16), and then mixed in an Erlenmeyer flask.

[0219] (7) Fermentation: The conical flasks were placed in a shaker at 28°C and 120 rpm for cultivation. The specific fermentation process is shown in Table 3-1.

[0220] (8) Sampling: On day 0, day 3 and day 4 of fermentation, approximately 45.0 mL of each sample was taken in a clean bench for determining the OD of the fermented tea vinegar. 600 pH, total acidity (TA), and sensory evaluation.

[0221] Table 3-1 Fermentation process parameters for different tea vinegars

[0222] 3.3 Experimental Results

[0223] In a three-strain fermentation system of yeast Y5-6, YF9-1 and lactic acid bacteria, the results showed that the system produced the best acid after 4 days of fermentation. Among them, the L-Y7-6 group had the highest total acid (Figure 4), the lowest pH (Figure 2), and the fastest growth (Figure 3).

[0224] 2. Sensory evaluation results

[0225] Sensory evaluation involved 15 participants. Each participant drank all five test samples in one round, with a 120-second rest between each sample. The samples were filtered between samples using water and Sudan biscuits. The evaluation focused on four aspects: aroma, taste, mouthfeel, and defects, culminating in a final overall preference rating. The overall preference rating used a 9-point scale: 1 point - extremely dislike; 5 points - neither dislike nor dislike; 9 points - extremely like. A score of 5.4 was generally considered satisfactory, and 7.2 points - excellent.

[0226] Overall, the fermented tea vinegar exhibited the highest flavor richness after 4 days of fermentation. Group L-Y7-6 showed the most abundant acidity and flavor, with a balanced overall taste. It also exhibited certain apricot, plum, and lemon-black tea flavors, achieving the highest sensory score (Figure 5). In the complex fermentation system, strains FZ022, XS001, and L-M23-6 produced a strong and pungent acetic acid flavor, failing to develop the characteristic flavor profiles of their strains, and were accompanied by unpleasant flavors such as astringency, bitterness, and fishiness.

[0227] Example 4. Fermented fruit juice

[0228] 4.1 Experimental Materials

[0229] Experimental strain: Lactobacillus plantarum L-Y7-6.

[0230] Laboratory supplies: pH meter (Mettler, Sz-standard); saccharimeter (Aito, RL-1); potentiometric titrator (Mettler, TITRATOR T50); full-wavelength microplate reader (Epoch2, equipment number 70000817); UV spectrophotometer (Shimadzu, Japan, 70001618); high-temperature steam sterilizer (Puwashi Co., Ltd., MLS-3781L-PC); incubator (Yonglian Bio, UL3L-12); analytical balance (Mettler, SP029); clean bench (equipment number YST-YJS-0025); centrifuge (5810R / Eppendorf, equipment number 70000218).

[0231] White grape juice, grapefruit juice, sugarcane juice, and MRS culture medium (Qingdao Haibo, batch number 20210603).

[0232] 4.2 Experimental Scheme

[0233] (1) Activation of bacterial strain: 1.0 mL (2.0%) of Lactobacillus plantarum L-Y7-6 preserved in glycerol tube was inoculated into 50 mL of MRS medium and incubated at 37°C for 24 h.

[0234] (2) Preparation of fermentation strain: Take 40.0 mL of the lactic acid bacteria solution prepared in step (1) into a 50.0 mL centrifuge tube in a clean bench, and then centrifuge at 6000 rpm for 15 min. After centrifugation, discard the supernatant, add about 45.0 mL of sterile RO water (water treated by reverse osmosis technology) to the centrifuge tube, resuspend the bacterial solution, centrifuge again at 6000 rpm for 15.0 min, discard the supernatant, and add 40.0 mL of sterile tea soup to make the seed culture.

[0235] (3) Preparation of fermentation base: Weigh a certain amount of fruit juice into a stainless steel beaker (as shown in Table 4-1), add water to a certain mass, and then stir evenly to prepare the fermentation base. Next, weigh 400.0g of fermentation base into an Erlenmeyer flask, and seal it with double-layer aluminum foil and sealing film.

[0236] (4) Sterilization: The treated fermentation substrate is pasteurized in an autoclave (70℃ for 15 min).

[0237] (5) Inoculation: After sterilization, cool to room temperature, inoculate the seed liquid obtained in step (2) at a concentration of 5% in the clean bench, and then mix it in an Erlenmeyer flask.

[0238] (6) Fermentation: The conical flasks were placed in a shaker at 28℃ and 120rpm for cultivation. The specific fermentation process is shown in Table 4-1.

[0239] (7) Sampling: On days 1, 2, and 3 of fermentation, approximately 45.0 mL of sample was taken in a clean bench for determining the sugar content (Brix%) and OD of the fermentation broth. 600 Total acidity and sensory evaluation.

[0240] Table 4-1

[0241] 4.3 Experimental Results

[0242] Figure 6-8 shows that *Lactobacillus plantarum* L-Y7-6 can ferment and grow normally in juices with different acidities and carbon sources, especially in white grape juice. In group 3 (white grape juice with 8.1% carbon source), the OD value reached 1.19 after 3 days of fermentation.

[0243] Sensory evaluation results showed that the fermented juice from white grapes after 3 days of fermentation had the highest sensory score. It retained some of the grape flavor and was rich in acidity, while also adding some lemony aroma, making it quite distinctive, as shown in Figure 9.

[0244] Example 5. Determination of antioxidant capacity and tyrosinase inhibitory capacity

[0245] 5.1 Experimental Materials

[0246] Experimental strains: see Table 5-1.

[0247] Table 5-1 Experimental strains

[0248] Laboratory supplies: Microplate reader, SPECTROstar Nano; Biochemical incubator, SPX-250B-Z; Vortex shaker, IKA VORTEX 3.

[0249] 5.2 Experimental Scheme

[0250] (1) Cell disruption: *Lactobacillus plantarum* L-Y7-6 and L-M23-6 were inoculated at 1% in MRS and cultured statically at 37°C for 24 h. Cells were disrupted using a cryogenic grinder, and the supernatant was collected by centrifugation for later use. (Other preparations have been completed previously.)

[0251] (2) Sampling: Take 6 μL of the sample from step (1) and determine the total antioxidant capacity (T-AOC), superoxide anion scavenging capacity, ABTS free radical scavenging capacity, DPPH free radical scavenging capacity, hydroxyl free radical scavenging capacity, and tyrosinase inhibition capacity. All the above determinations were performed using commercially available kits and in accordance with the instructions of each kit. The positive control sample used was vitamin C, and the specific concentration is shown in Table 5-2.

[0252] 5.3 Experimental Results

[0253] Table 5-2 Results of Antioxidant and Tyrosinase Inhibition Properties Tests

[0254] Compared to *Lactobacillus plantarum* L-M23-6, *Lactobacillus plantarum* L-Y7-6 showed significantly superior antioxidant properties and tyrosinase inhibition rate, with a stronger overall antioxidant capacity. Specifically, the tyrosinase inhibition rate of L-Y7-6 was 1.65 times that of L-M23-6 at 3 minutes and 1.35 times that of L-M23-6 at 15 minutes. Notably, in terms of total antioxidant capacity, L-Y7-6 was significantly higher than the positive control sample (0.5 mmol / L Vc). This indicates that *Lactobacillus plantarum* L-Y7-6 possesses certain antioxidant capabilities and has potential applications in the cosmetics field. Furthermore, it also has the potential to improve the antioxidant capacity, nutritional value, and preservation performance of fermented fruit juices, fermented teas, and fermented tea vinegars.

[0255] Example 6. Antioxidant Study of L-Y7-6 Zebrafish

[0256] In this embodiment, the antioxidant efficacy of culture supernatants and lysates of three Lactobacillus plantarum strains L-Y7-6, XS002, and XS004 was studied based on a zebrafish antioxidant model.

[0257] 1. Model Building Principles

[0258] Oxidative stress is clearly defined as a condition caused by an imbalance in the cellular redox state. In living cells, prooxidant species originate from oxygen (ROS; reactive oxygen species) or nitrogen (RNS; reactive nitrogen species). ROS include hydroxyl radicals (.OH), superoxide anions (O2-), and hydrogen peroxide (H2O2). Primary RNS is nitrous oxide (NO). High levels of reactive oxygen species (ROS) can lead to a shift in the cellular redox state towards oxidative stress. This condition results in the oxidation of molecules (lipids, DNA, proteins) and ultimately cell death.

[0259] Zebrafish, as a model organism, possesses a complete in vivo system. Some toxicological studies have used zebrafish to assess the in vivo effects of chemicals on redox homeostasis, indicating that this vertebrate is suitable as an animal model in drug discovery and oxidative stress. Tert-Butyl hydroperoxide (t-BHP) is an organic peroxide. As a derivative of hydrogen peroxide, it rapidly decomposes within cells, generating free radicals such as hydroxyl radicals (·OH) and peroxide groups (ROO·). These reactive oxygen species (ROS) can cause oxidative damage to various intracellular biomolecules such as lipids, proteins, and DNA. Previous studies have shown that t-BHP can induce oxidative stress in zebrafish.

[0260] 2. Materials and Methods

[0261] 2.1 Materials

[0262] 2.1.1 Experimental Zebrafish

[0263] Wild-type zebrafish: AB strain 3dpf.

[0264] 2.1.2 Sample Information

[0265] A total of 12 samples were evaluated. Detailed sample information is shown in Table 6-1 below. SH refers to the culture supernatant of the bacterial strain, and RB refers to the lysate of the bacterial strain. The supernatant was prepared by centrifuging the three cultured bacterial suspensions separately to obtain the supernatant for later use. The lysate was prepared by disrupting the cells using a high-pressure cell disruptor at a pressure of 1000-1400 ber, centrifuging to obtain the supernatant, and then reserving the lysate for later use.

[0266] Table 6-1 Test Sample Information Table

[0267] 2.1.3 Experimental Apparatus

[0268] 2.1.4 Experimental Reagents and Consumables

[0269] 2.1.5 Preparation of Experimental Reagents

[0270] 1) 50×E3 buffer stock solution: Accurately weigh 14.60g NaCl, 0.65g KCl, and 2.20g CaCl₂. 2， 6.12g MgSO4·7H2O was placed in a 1L volumetric flask, diluted to volume with ultrapure water, and magnetically stirred for 48h. It was then stored at room temperature for later use.

[0271] 2) E3 buffer working solution: Take 100mL of 50×E3 buffer, dilute to 5L with ultrapure water, shake well and set aside at room temperature;

[0272] 3) Molding agent t-BHP (3mM): The molding agent stock solution is a liquid with a mass fraction of 70%, and should be prepared and used immediately.

[0273] 4) Preparation of positive control table gallocatechin gallate (EGCG) stock solution (12.5 mg / mL): Accurately weigh 12.5 mg ascorbic acid into a 1.5 mL centrifuge tube, add 1 mL of ultrapure water to the centrifuge tube, vortex and sonicate until completely dissolved, and use immediately after preparation;

[0274] 5) Test Samples: All submitted samples were liquids, and were administered at six concentration gradients using E3 buffer working solution diluted fourfold. The dilution method was as follows: 4 mL of sample solution was added to a 6-well plate, and 1 mL of sample solution was aspirated and added to a 6-well plate containing 3 mL of E3 buffer. The mixture was thoroughly mixed with a pipette, and six concentrations were administered sequentially: 98%, 25%, 6.25%, 1.56%, 0.39%, and 0.10%.

[0275] 2.2 Experimental Methods

[0276] 2.2.1 Determination of Maximum Tolerable Concentration (MTC)

[0277] 1) Take a six-well cell culture plate and set up a blank control group, a model control group, a positive control group, and a sample group. Select zebrafish juveniles with normal development and 3 dpf and put them into a six-well cell culture plate, 20 fish per well. Incubate in a biochemical incubator at (28.5℃±1.0℃) for 24 hours. At this time, the zebrafish are 4 dpf.

[0278] 2) Take a new six-well cell culture plate, add 3 mL of E3 Buffer working solution to the blank control group, and add 3 mL of modeling agent t-BHP with a final concentration of 3 mM to the six-well plates of the other groups. Place the zebrafish of each group in the corresponding group and incubate in a (28.5℃±1.0℃) biochemical incubator for 1 h.

[0279] 3) The concentration group of zebrafish juveniles that showed death (no heartbeat) and other toxic effects (pericardial edema, trunk bending, no response to mechanical stimulation, unclear muscle texture, etc.) was defined as MTC.

[0280] 2.2.2 Efficacy Test

[0281] Repeat the experimental steps 1-2 in 2.2.1.

[0282] After modeling, 15 zebrafish were collected from each group and stored in labeled 1.5 mL centrifuge tubes. The tubes were dried, and 500 μL of E3 Buffer and 3 zirconium beads were added. The tubes were then ground in a tissue homogenizer for 1 min 30 s until the tissue was completely homogenized. The tubes were centrifuged at 4000 rpm for 5 min, the liquid was aspirated, and 500 μL of DPBS containing DCFH-DA (10 μM) and Hoechst (1:4000) probes was added. After vortexing, the tubes were incubated at 37°C in the dark for 20 min, centrifuged at 4000 rpm for 5 min, the liquid was aspirated, and 300 μL of DPBS was added. The mixture was vortexed, and 100 μL of the suspension was added to a 96-well black plate. The fluorescence intensity at excitation light 485 nm and emission light 528 nm (DCFH-DA) and excitation light 350 nm and emission light 461 nm (Hoechst) was measured using a microplate reader.

[0283] 2.3 Experimental Results and Statistics

[0284] The obtained fluorescence intensity values ​​were used to calculate the ROS intensity, with OD = OD DCFH-DA / OD Hoechst The fluorescence intensity was used as the calibrated value. Analysis of variance was performed on the data using Graphpad Prism 10.0 software, with the model control group as the standard. The fluorescence intensity of each experimental group was compared, and P ≤ 0.05 indicated a significant difference.

[0285] Compared with the blank control group, the fluorescence intensity of the model control group was significantly increased (P≤0.05), while the fluorescence intensity of the test substance was significantly decreased (P≤0.05) compared with the model control group, indicating that the test substance has antioxidant effects at this concentration.

[0286] 3. Experimental Results

[0287] 3.1 MTC Results

[0288] Table 6-2 MTC Results for Each Sample

[0289] 3.2 Results of Antioxidant Efficacy Evaluation Experiment

[0290] Based on the preliminary experimental results, the antioxidant efficacy of the samples was evaluated under MTC conditions.

[0291] Table 6-3 Results of antioxidant effects Note: ### indicates P≤0.001 between the model group and the control group; *** indicates P≤0.001, and * indicates P≤0.05 between the sample group and the model group.

[0292] Experimental conclusions

[0293] Compared with the two identical strains, L-Y7-6 exhibited lower levels of reactive oxygen species (ROS) and stronger antioxidant activity in both its supernatant and lysate. Specifically, the antioxidant rate of L-Y7-6 was significantly different from that of the model group (P≤0.05*), while neither of the two identical strains showed a significant difference compared to the model group.

[0294] Furthermore, according to MTC test results, L-Y7-6 exerts its antioxidant effects without causing toxic side effects on zebrafish. Therefore, L-Y7-6 has high potential for antioxidant applications.

[0295] Example 7. Study on the laxative effect of L-Y7-6

[0296] 1. Experimental Objective

[0297] This embodiment establishes a mouse small intestinal peristalsis inhibition model to evaluate the effects of different samples on mouse intestinal peristalsis.

[0298] 2. Experimental Principle

[0299] Constipation is one of the most common and complex digestive disorders in clinical practice, and also a complication of other diseases. With changes in people's dietary structure and lifestyle, the incidence of constipation has risen abnormally. Constipation can cause a variety of symptoms, including abdominal distension, fatigue, weight loss, loss of appetite, dull complexion, age spots, halitosis, and slowed reactions, resulting in dark, dry stools. Patients also experience discomfort and pain, severely impacting their quality of life.

[0300] A mouse model of small intestinal peristalsis inhibition was established by oral gavage administration of loperamide, a drug that inhibits intestinal peristalsis. Subsequently, the mice were given ink, and the ink propulsion rate in the small intestine was calculated over a certain period of time to assess the gastrointestinal function of the model mice.

[0301] 3 Materials and Methods

[0302] 3.1 Materials

[0303] 3.1.1 Laboratory Animals

[0304] Male C57 mice, 6 - 8 weeks old, weighing 20 - 22 g, 71 in number, were purchased from Shanghai Slake Experimental Animal Co., Ltd. (License: SCXK(Shanghai)2022 - 0004), and the certificate number was 20220004079115. The animal room was maintained under a strict 12 / 12 h day / night cycle, and the mice were provided with sufficient regular feed and water. All animal experiments were approved by the Animal Ethics Committee.

[0305] Experimental grouping: The experiment set up 3 sample groups, one blank control group and one model control group. The blank control group and the model control group were given distilled water by the same route, with 13 - 14 mice in each group. The test samples were administered for 28 days.

[0306] 3.1.2 Experimental reagents and consumables

[0307] 3.1.3 Preparation of experimental reagents

[0308] Preparation of ink: Accurately weigh 50 g of gum arabic, add 400 mL of water, boil until the solution is transparent, weigh 25 g of activated carbon (powdered) and add it to the above solution, boil three times, after the solution cools, add water to make the volume up to 500 mL, store at 4 °C in the refrigerator, and shake well by ultrasonic heating before use.

[0309] Preparation of loperamide hydrochloride solution: Take 4 mg of loperamide, add pure water to 20 mL, dissolve and mix well for standby. [[

[0310] Preparation of bacterial powder: Wet bacterial cells were obtained through large - scale cultivation. Using a composite protective agent formula with skim milk, fructooligosaccharides and sodium glutamate as the core, after pre - freezing and freeze - drying for up to 56 hours, the bacterial powder was successfully prepared. Calculate the survival rate of the bacterial powder, and configure the specific dosage according to the bacterial viability, and prepare a suspension with the corresponding normal saline.

[0311] 3.2 Establishment of the model

[0312] After all mice were adaptively fed for 1 week, they were randomly grouped according to body weight, with 13 - 14 mice in each group, a total of 5 groups. They were respectively sample groups 1 - 3 (RC 8 - 1—RC 8 - 3), blank control group (C) and model control group (M), with 13 - 14 mice in each group. The samples and dosing information are as follows in the table. The test samples were administered for 28 days, and the mice in each group were fasted for 16 hours without water deprivation. The model control group and the 4 sample groups were intragastrically administered loperamide (4 mg / kg BW), and the blank control group was given distilled water to establish a mouse small intestine peristalsis inhibition model.

[0313] 3.3 Detection of small intestine motor function

[0314] Mice were fasted for 16 hours but allowed free access to water. The following morning, the model group and three sample groups were administered loperamide 4 mg / (kg·Bw) by gavage, while the blank control group received an equal volume of pure water. The gavage volume for all groups was 50 mL / (kg·Bw). 0.5 h after loperamide administration, each group was administered an ink mixture by gavage, with a gavage volume of 50 mL / (kg·Bw). Immediately after 25 min, the animals were euthanized by cervical dislocation. The abdominal cavity was opened, the mesentery was separated, and the intestinal segment from the pylorus to the ileocecal junction was cut off. The segment was placed on a tray, and the small intestine was gently stretched into a straight line. The length of the segment was measured as the "total length of the small intestine," and the distance from the pylorus to the ink front was measured as the "ink propulsion length."

[0315] Calculate the ink propulsion rate using the following formula:

[0316] 3.4 Statistical Processing

[0317] Statistical analysis was performed using GraphPad software. Data are expressed as mean ± standard deviation (Mean ± SD). One-way Anova analysis was used for comparisons among multiple groups, with P < 0.05 considered statistically significant and P < 0.01 considered highly significant. Nonparametric tests were performed on non-normal or unequal variance data.

[0318] 4. Experimental Results

[0319] 4.1 Analysis of Small Intestinal Motility

[0320] At the end of the experiment, the length of the intestinal segment was measured as the "total length of the small intestine", and the distance from the pylorus to the ink front was measured as the "ink propulsion length". The ink propulsion rate was calculated to assess the motility of the small intestine.

[0321] 5. Experimental Conclusions

[0322] The ink propulsion rate of L-Y7-6 was significantly higher than that of the model group, indicating that it can effectively alleviate the inhibition of small intestinal peristalsis caused by loperamide and has a laxative effect that promotes intestinal peristalsis.

[0323] Example 8. Evaluation of L-Y7-6 carbon clearance efficiency

[0324] 1. Experimental Objective

[0325] The effects of different samples on mouse monocyte-macrophage function were evaluated by establishing a mouse carbon clearance model.

[0326] 2. Experimental Principle

[0327] Within a certain range, the rate at which carbon particles are cleared from the body exhibits an exponential relationship with blood carbon concentration.

[0328] Taking the logarithm of blood carbon concentration as the ordinate and time as the abscissa, there is a linear relationship between the two. The slope (K) of this straight line can represent the phagocytosis rate. The weights of the animal liver and spleen affect the phagocytosis rate, which is generally represented by the corrected phagocytosis index a.

[0329] 3 Materials and Methods

[0330] 3.1 Materials

[0331] 3.1.1 Experimental Animals

[0332] Male ICR mice, 6 - 8 weeks old, weighing 22 - 25 g, 50 in number, were purchased from Shanghai Slake Experimental Animal Co., Ltd. (License: SCXK(Shanghai)2022 - 0004), and the certificate number was 20220004083724. The animal house was strictly in a 12 / 12 h day / night cycle, and sufficient regular feed and water were provided. All animal experiments were approved by the Animal Ethics Committee.

[0333] Experimental grouping: The experiment was set up with 3 sample groups, one positive control group and one model control group. The negative control group and the model control group were given distilled water by the same route, with 10 mice in each group. The test samples were administered for 30 days.

[0334] 3.1.2 Experimental Reagents and Consumables

[0335] 3.1.3 Preparation of Experimental Reagents

[0336] Preparation of India ink: Dilute the original India ink stock solution 3 times with normal saline.

[0337] Na2CO3 solution: Take 0.1 g of Na2CO3 and add distilled water to 100 mL.

[0338] Levamisole solution (25 mg / kg): Take one tablet and dissolve it in 20 mL of RO water.

[0339] Preparation of bacterial powder: Wet bacterial cells were obtained through large - scale cultivation. Using a composite protective agent formula with skim milk, fructooligosaccharides and sodium glutamate as the core, after pre - freezing and freeze - drying for up to 56 hours, the bacterial powder was successfully prepared, and the survival rate of the bacterial powder was calculated. According to the bacterial viability, the specific dosage for administration was configured and made into a suspension with the corresponding normal saline.

[0340] 3.2 Establishment of the Model

[0341] After one week of acclimatization, all mice were randomly divided into five groups of 10 mice each, based on their body weight. These groups were: sample groups 1-3 (TKQ2-1-TKQ2-3), a blank control group (C), and a positive control group (levamisole tablets), with 10 mice in each group. Sample and administration information is shown in the table below. The test samples were administered for 30 days, while the positive control group received the medication for the last three days. Mice in all groups were fasted for 16 hours after the last administration, but allowed free access to water. The blank control group, positive control group, and the three sample groups were injected intravenously with 10 mL / kg of diluted ink.

[0342] 3.3 Detection of carbon clearance capacity in mice

[0343] Two minutes (t2) and ten minutes (t1) after ink injection, 20 μL of blood was drawn from the internal canthal venous plexus and immediately added to 2 mL of 0.1% Na2CO3 solution. The optical density (OD) was measured at 600 nm using a 721 spectrophotometer or an ELISA reader.

[0344] The mice were euthanized, and their livers and spleens were removed. The blood on the surface of the organs was absorbed with filter paper, and the organs were weighed separately.

[0345] The phagocytic index represents the carbon clearance capacity of mice. The phagocytic index 'a' is calculated using the following formula. A significantly higher phagocytic index in the test sample group compared to the control group indicates a positive result for this experiment.

[0346] Calculate the phagocytic index using the following formula:

[0347] 3.4 Statistical Processing

[0348] Statistical analysis was performed using GraphPad software. Data are expressed as mean ± standard deviation (Mean ± SD). One-way Anova analysis was used for comparisons among multiple groups, with P < 0.05 considered statistically significant and P < 0.01 considered highly significant. Nonparametric tests were performed on non-normal or unequal variance data.

[0349] 4 Experimental Results

[0350] The phagocytic index of L-Y7-6 was significantly higher than that of the control group, indicating that it can enhance the non-specific immune function of mice and promote the clearance of carbon particles.

[0351] Although specific embodiments of the invention have been described in detail, those skilled in the art will understand that various modifications and variations can be made to the details based on all the published teachings, and all such changes are within the scope of protection of the invention. The entire scope of the invention is given by the appended claims and any equivalents thereof.

Claims

1. A species of Lactiplantibacillus plantarum, deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 31269.

2. The *Lactobacillus plantarum* according to claim 1, having one or more of the following characteristics: (1) Having a 16S rDNA sequence that is 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9%, 99.95% or 100% identical to SEQ ID NO:1; (2) The plant lactobacillus has the ability to produce more than 1 g / kg (e.g., more than 1.2 g / kg, more than 1.5 g / kg, more than 1.8 g / kg, more than 2.1 g / kg) of total acid under effective culture conditions; (3) The plant lactobacillus can reduce the pH of the fermentation product (e.g., pH reduction of 0.6 or more); (4) The plant lactobacillus can produce a variety of volatile aroma components under effective culture conditions, and the flavor is selected from fruity flavor, floral flavor, milky flavor, woody flavor, or any combination thereof; (5) The plant lactobacillus can resist oxidation and / or inhibit tyrosinase.

3. A composition comprising *Lactobacillus plantarum* as described in claim 1 or 2; Preferably, the composition further comprises microorganisms selected from bacteria, fungi (e.g., yeast), or any combination thereof; preferably, the microorganisms are probiotics.

4. The composition of claim 3, wherein, The bacteria are selected from the genera *Lactobacillus*, *C.*, *Bifidobacterium*, *Lactobacillus mucosae*, *Lactobacillus*, *L. assemblica*, *Lactobacillus spp.*, *Streptococcus*, *Lactococcus*, *Propionibacterium*, *Leuconostoc*, *Pediococcus*, *Weizmannella*, *Azoococcus*, *Staphylococcus*, *Bacillus*, *Acetobacter*, *Coprinus*, *Gluconobacterium*, *Gluconobacterium*, or any combination thereof; Preferably, the bacteria of the genus *Lactobacillus* are selected from: *Lactiplantibacillus plantarum*, *Lactobacillus casei*, *Lactobacillus paracasei*, *Lactobacillus brevis*, *Lactobacillus pentosus*, *Lactobacillus crispatus*, *Lactobacillus rhamnosus*, or any combination thereof; Preferably, the yeast is selected from Zygosaccharomyces bisporus, Dekkera Bruxellensis, Saccharomyces cerevisiae, Saccharomyces boulardii, Kluyveromyces marxianus, or any combination thereof.

5. The composition of claim 3 or 4, wherein the composition comprises *Lactobacillus plantarum* as described in claim 1 or 2, and *Diplostomum bifidum* and / or *Dacronella brusselii*. Preferably, the *Bacillus zygosacchariformis* is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 31264; Preferably, the *Dacronis brusselsii* is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 28050; Preferably, the ratio of *Zygosaccharomyces bisporus*, *Dacronis brusselsii*, and *Lactobacillus plantarum* in the composition is 1-10:1-10:10-20 (e.g., 1:8:16, 1:8:10-20, 1-10:8:16, 1:1-10:16, 1:1-10:10-20, 1-10:8:10-20, 1-10:1-10:16). Preferably, the ratio of *Dictyophora indicum*, *Dictyophora brusselii*, and *Lactobacillus plantarum* in the composition is 1:8:16; Preferably, the composition is used for fermentation; Preferably, the composition further includes additional additives; Preferably, the additional additives are selected from other nutrients (e.g., carbon sources, nitrogen sources, vitamins, minerals, trace elements, growth factors, or any combination thereof).

6. A culture comprising *Lactobacillus plantarum* as described in claim 1 or 2, or a composition as described in any one of claims 3-5; Preferably, the culture further includes additional additives; Preferably, the additional additives are selected from other nutrients (e.g., carbon sources, nitrogen sources, vitamins, minerals, trace elements, growth factors, or any combination thereof).

7. The culture of claim 6, wherein, The carbon source is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof; Preferably, the nitrogen source is selected from ammonia, peptone, yeast extract, soybean hydrolysate, milk, or any combination thereof.

8. The culture according to claim 6 or 7, wherein, The additional additives also include tea and / or fruit juice; Preferably, the tea infusion is selected from black tea infusion, green tea infusion, flower tea infusion, Longjing tea infusion, white tea infusion, Lao Ying tea infusion, oolong tea infusion, barley tea infusion, purple leaf tea infusion, or any combination thereof; preferably, the tea infusion is black tea infusion. Preferably, the culture is a solid, liquid, or semi-solid; Preferably, the culture further comprises a cell-free culture filtrate of the *Lactobacillus plantarum*.

9. The culture according to any one of claims 6-8, wherein, The culture also contains derivatives of Lactobacillus plantarum; Preferably, the derivative is selected from metabolites, supernatant of Lactobacillus plantarum, lysate of Lactobacillus plantarum, or any combination thereof.

10. A food product or dietary supplement comprising Lactobacillus plantarum as described in claim 1 or 2, or a composition as described in any one of claims 3-5, or a culture as described in any one of claims 6-9; Preferably, the food product or dietary supplement is a beverage; Preferably, the food product or dietary supplement is selected from tea drinks, solid beverages, or fruit juice drinks; Preferably, the food product or dietary supplement contains dairy products (e.g., yogurt, flavored fermented milk, lactic acid bacteria beverages, cheese).

11. A fermentation method comprising: inoculating a tea infusion with *Lactobacillus plantarum* as described in claim 1 or 2, or a composition as described in any one of claims 3-5, or a culture as described in any one of claims 6-9; optionally, culturing the *Lactobacillus plantarum* in the inoculated tea infusion.

12. A fermentation product obtained by fermentation of *Lactobacillus plantarum* as described in claim 1 or 2, or a composition as described in any one of claims 3-5, or a culture as described in any one of claims 6-9, or obtained by the fermentation method as described in claim 11; Preferably, the fermented product is fermented tea, fermented tea vinegar, fermented fruit juice, fermented dairy products, or any combination thereof; Preferably, the fermentation product further includes additional additives; Preferably, the additional additives are selected from sweeteners, colorings, preservatives, antioxidants, acidity regulators, nutritional fortifiers, or any combination thereof.

13. The fermented product of claim 12, wherein the fermented tea has one or more of the following characteristics: (1) The fermented tea is prepared by fermentation using Lactobacillus plantarum as described in claim 1 or 2, or it is obtained by the fermentation method described in claim 11; (2) The tea is selected from black tea, green tea, flower tea, Longjing tea, white tea, Lao Ying tea, oolong tea, barley tea, or any combination thereof; preferably, the tea is black tea; Preferably, the tea content is 0.2%-1.6% (m / v) (e.g., 0.2%, 0.4%, 0.8%, 1.2%, 1.6%); preferably, the tea content is 0.8% (m / v). (3) The total sugar content (Brix%) of fermentation is 2%-8% (m / v) (e.g., 2%, 4%, 6%, 8%); Preferably, the total sugar content (Brix%) of the fermentation is 4% (m / v); (4) The carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof; Preferably, the carbon source for fermentation is selected from sucrose, glucose, grapefruit juice, or any combination thereof; Preferably, the carbon source for fermentation is sucrose, glucose and grapefruit juice in a ratio of 4:2:3; Preferably, the carbon source for fermentation is sucrose, glucose, and grapefruit juice, with the addition amounts being 2% sucrose, 1% glucose, and 1.5% grapefruit juice; (5) The fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days); Preferably, the fermentation time is 3 days; (6) The fermented tea has a lemon aroma.

14. The fermented product according to claim 12 or 13, wherein the fermented tea vinegar has one or more characteristics selected from the following: (1) The fermented tea vinegar is prepared by fermentation of a yeast containing zygosacchariformis, Dioscorea brussels yeast and Lactobacillus plantarum as described in claim 1 or 2, or it is obtained by the fermentation method described in claim 11; (2) The pH value of the tea vinegar is less than 4 (for example, pH value less than 3.6, pH value less than 3.2); (3) The tea in the tea vinegar is selected from black tea, green tea, flower tea, Longjing tea, white tea, Lao Ying tea, oolong tea, barley tea, or any combination thereof; preferably, the tea in the tea vinegar is black tea; Preferably, the tea content is 0.2%-1.6% (m / v) (e.g., 0.2%, 0.4%, 0.8%, 1.2%, 1.6%); preferably, the tea content is 0.8% (m / v). (4) The total sugar content (Brix%) of fermentation is 2%-8% (m / v) (e.g., 2%, 4%, 6%, 8%); Preferably, the total sugar content (Brix%) of the fermentation is 4% (m / v); (5) The carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof; Preferably, the carbon source for fermentation is selected from sucrose, glucose, grapefruit juice, or any combination thereof; Preferably, the ratio of sucrose, glucose, and grapefruit juice as carbon sources for fermentation is 4:2:3; Preferably, the amount of sucrose, glucose, and grapefruit juice added as carbon sources for fermentation is 2% sucrose, 1% glucose, and 1.5% grapefruit juice; (6) The fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days); Preferably, the fermentation time is 3 days.

15. The fermented product of claim 14, wherein the fermented tea vinegar is prepared by fermentation comprising *Zygosaccharomyces cerevisiae*, *Dacronis brusselsii*, and *Lactobacillus plantarum* as described in claim 1 or 2; Preferably, the *Bacillus zygosacchariformis* is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 31264; Preferably, the *Dacronis brusselsii* is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 28050; Preferably, the ratio of *Zygosaccharomyces bisporus*, *Dacronis brusselsii*, and *Lactobacillus plantarum* is 1-10:1-10:10-20 (e.g., 1:8:16, 1:8:10-20, 1-10:8:16, 1:1-10:16, 1:1-10:10-20, 1-10:8:10-20, 1-10:1-10:16). Preferably, the ratio of the two-spore conjugated yeast, the Brussels yeast, and the plant lactobacillus is 1:8:

16.

16. The fermented product according to any one of claims 12-15, wherein the fermented juice has one or more characteristics selected from the following: (1) The fermented fruit juice is prepared by fermentation using Lactobacillus plantarum as described in claim 1 or 2, or it is obtained by the fermentation method described in claim 11; (2) The carbon source for fermentation is selected from sucrose, glucose, fructose, maltose, lactose, grapefruit juice, grape juice, sugarcane juice, or any combination thereof; preferably, the carbon source for fermentation is white grape juice; Preferably, the carbon source for fermentation is 5%-10% white grape juice (e.g., 8.1%); (3) The fermentation time is 1-4 days (e.g., 1 day, 2 days, 3 days, 4 days); Preferably, the fermentation time is 3 days; (4) The fermentation temperature is 23-32℃ (e.g., 24℃, 26℃, 28℃, 30℃).

17. Use of the *Lactobacillus plantarum* according to claim 1 or 2, or the composition according to any one of claims 3-5, or the culture according to any one of claims 6-9, in fermentation; Preferably, the fermentation is used to prepare fermented tea, fermented tea vinegar, fermented fruit juice, fermented dairy products, or any combination thereof.

18. Use of the *Lactobacillus plantarum* according to claim 1 or 2, or the composition according to any one of claims 3-5, or the culture according to any one of claims 6-9, in the preparation of antioxidants; Preferably, the antioxidant is capable of inhibiting tyrosinase; Preferably, the antioxidant is capable of scavenging superoxide anions, scavenging ABTS free radicals, scavenging DPPH free radicals, scavenging hydroxyl free radicals, and / or inhibiting superoxide dismutase (SOD).

19. Use of the *Lactobacillus plantarum* according to claim 1 or 2, or the composition according to any one of claims 3-5, or the culture according to any one of claims 6-9 in the preparation of food products or dietary supplements; Preferably, the food product or dietary supplement is a beverage; Preferably, the food product or dietary supplement is selected from tea drinks, solid beverages, or fruit juice drinks; Preferably, the food product or dietary supplement contains dairy products (e.g., yogurt, flavored fermented milk, lactic acid bacteria beverages, cheese).

20. Use of the *Lactobacillus plantarum* of claim 1 or 2, or the composition of any one of claims 3-5, or the culture of any one of claims 6-9, in the preparation of a medicament for the prevention and / or treatment of constipation in a subject, or for enhancing the immune function (e.g., nonspecific immunity) of a subject.

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