Phytobacterium plantarum TLP03, preparation, application and product

Abstract

The invention relates to the technical field of microbial fermentation, in particular to lactobacillus plantarum TLP03, a preparation, application and a product, the lactobacillus plantarum TLP03 is preserved in the China Center for Type Culture Collection, the preservation name of the lactobacillus plantarum TLP03 is TLP03, and the preservation number of the lactobacillus plantarum TLP03 is CCTCC NO: M20242276. The plant lactobacillus TLP03 not only has low pH resistance and salt resistance, but also has high yield of lactic acid, acetic acid, glutamic acid and hexanal, and when the plant lactobacillus TLP03 is applied to the fermentation process of pickled Chinese cabbages, the taste of the prepared pickled Chinese cabbages can be improved.

Description

Technical Field

[0001] The present invention relates to the technical field of microbial fermentation, and specifically relates to a Lactiplantibacillus plantarum TLP03, a preparation and application thereof, and a product. Background Art

[0002] As a traditional fermented vegetable product in China, pickled Chinese cabbage is deeply loved by the public because of its unique taste and rich nutritional value. Especially in the southwestern region of China, pickled Chinese cabbage is almost a necessity in every household.

[0003] The preparation process of pickled Chinese cabbage mainly includes processes such as salting in the pool, secondary pickling in the altar, packaging, and sterilization. Among them, secondary pickling in the altar is the fermentation process. Usually, the pickled vegetable embryos after salting are washed and then transferred into the altar, and brine with a salinity of 8-13% is added for fermentation to obtain pickled Chinese cabbage. That is, the fermentation method of secondary pickling in the altar is mainly natural fermentation. In the brine immersion environment, various microorganisms attached to the surface of vegetables rapidly reproduce using nutrients such as sugars in the raw materials. Sourness is the most basic element in the taste of pickled Chinese cabbage, mainly derived from organic acids such as lactic acid produced by the metabolism of lactic acid bacteria.

[0004] As a generally recognized as safe (GRAS) strain, lactic acid bacteria have attracted much attention due to their wide application as biological additives in foods. Lactiplantibacillus plantarum ( Lactiplantibacillus plantarum ) is a common lactic acid bacterium and is a member of the genus Lactiplantibacillus ( Lactobacillus ), and its main characteristics include acid resistance, salt tolerance, lactic acid fermentation, and probiotic effects.

[0005] Therefore, Lactiplantibacillus plantarum can adapt to the low pH and high salt environments in the pickled Chinese cabbage fermentation process. However, for Lactiplantibacillus plantarum used in the pickled Chinese cabbage fermentation process, it is not only required to have low pH resistance and salt tolerance, but also to have properties such as low temperature resistance and high production of organic acid flavor substances. Summary of the Invention

[0006] The purpose of the present invention is to provide a Lactiplantibacillus plantarum that has low pH resistance and salt tolerance, and also has high production of lactic acid, acetic acid, glutamic acid, and hexanal.

[0007] In addition, the present invention also provides the application of the above Lactiplantibacillus plantarum, as well as a preparation, pickled Chinese cabbage, a preparation method of pickled Chinese cabbage, and a pickled Chinese cabbage-based pickled fish seasoning packet prepared based on it.

[0008] Using the Lactiplantibacillus plantarum 3 of the present invention for pickled Chinese cabbage fermentation can make the prepared pickled Chinese cabbage taste better.

[0009] The present invention is achieved through the following technical solutions: A Lactiplantibacillus plantarum, the Lactiplantibacillus plantarum ( Lactiplantibacillus plantarum)TLP03 is deposited in the China Center for Type Culture Collection under the deposit name of TLP03 and the deposit number of CCTCC NO: M20242276.

[0010] The original intention of the present invention is as follows: First, screen a strain of lactic acid bacteria that can highly produce substances improving the taste of pickled Chinese cabbage and can tolerate the low pH and high salt environment during the pickled Chinese cabbage fermentation process.

[0011] Second, apply the selected strain to the pickled Chinese cabbage fermentation, so that the prepared pickled Chinese cabbage contains relatively high levels of lactic acid, acetic acid, glutamic acid and hexanal, to improve the taste of pickled Chinese cabbage and provide new beneficial microbial fermentation strains for the pickled Chinese cabbage industry.

[0012] The deposit date of the above Lactiplantibacillus plantarum TLP03 is October 21, 2024, the deposit address is Wuhan University, Wuhan, China, Zip Code: 430072, and the taxonomic name is: Lactiplantibacillus plantarum .

[0013] The Lactiplantibacillus plantarum TLP03 of the present invention is isolated from the water sample of large-leaf pickled Chinese cabbage. After detection, it has the properties of tolerating low pH, salt, and highly producing lactic acid, acetic acid, glutamic acid and hexanal. Applying it to the pickled Chinese cabbage fermentation process can increase the contents of lactic acid, acetic acid, glutamic acid and hexanal in pickled Chinese cabbage, thereby improving the taste of pickled Chinese cabbage.

[0014] A preparation containing Lactiplantibacillus plantarum, and the form of the preparation includes bacterial agent, bacterial sludge, bacterial powder or bacterial liquid.

[0015] Application of Lactiplantibacillus plantarum in the pickled Chinese cabbage fermentation process.

[0016] In a preferred manner, first prepare Lactiplantibacillus plantarum TLP03 into bacterial sludge or bacterial liquid, and then inoculate the bacterial sludge or bacterial liquid into the pickled Chinese cabbage soaked in brine for fermentation.

[0017] In a preferred manner, the inoculation amount of the bacterial liquid is (8 - 12) g / kg; the inoculation amount of the bacterial sludge is (0.5 - 1.5) g / kg.

[0018] In a preferred manner, the inoculation amount of the bacterial liquid is (10 - 12) g / kg; the inoculation amount of the bacterial sludge is (1.0 - 1.5) g / kg.

[0019] In a preferred manner, the inoculation amount of the bacterial liquid is 10 g / kg; the inoculation amount of the bacterial sludge is 1.0 g / kg.

[0020] Specifically, the preparation process of the bacterial liquid is as follows: Inoculate Lactiplantibacillus plantarum TLP03 into MRS broth medium and activate it at 35 - 39 °C for 20 - 26 h to obtain a seed liquid; Inoculate the seed liquid into another MRS broth medium at an inoculation amount of 1-5% (v / v), and activate it at 35-39 °C for 20-26 h to obtain a fermentation broth, that is, a bacterial liquid. The preparation process of the bacterial sludge is as follows: Centrifuge the bacterial liquid at a centrifugation temperature of 2-6 °C, a centrifugation speed of 6000-10000 rpm, and a centrifugation time of 6-10 min. After centrifugation, collect the lower-layer bacterial precipitate to obtain the bacterial sludge.

[0021] Specifically, the composition of the MRS broth medium is: peptone 10.0 g / L, beef powder 5.0 g / L, glucose 20.0 g / L, yeast powder 4.0 g / L, sodium acetate 5.0 g / L, dipotassium hydrogen phosphate 2.0 g / L, magnesium sulfate 0.2 g / L, ammonium citrate 2.0 g / L, manganese sulfate 0.05 g / L, Tween 80 1 mL / L.

[0022] The application of the above preparation in the process of pickled Chinese cabbage fermentation.

[0023] A product, including pickled Chinese cabbage or a compound condiment containing pickled Chinese cabbage; The pickled Chinese cabbage is prepared by fermentation with Lactiplantibacillus plantarum TLP03 or the above-mentioned bacterial agent; The compound condiment includes pickled Chinese cabbage fish seasoning, pickled Chinese cabbage hot pot base material or pickled Chinese cabbage rice noodle seasoning. The compound condiment contains pickled Chinese cabbage. For example: the pickled Chinese cabbage fish seasoning includes a pickled Chinese cabbage packet, a seasoning packet and a pickled fish material packet, and the pickled Chinese cabbage packet contains the above-mentioned pickled Chinese cabbage. Specifically, the pickled Chinese cabbage packet includes a packaging bag, and the packaging bag contains pickled Chinese cabbage and an aqueous solution, and the solid content of the pickled Chinese cabbage packet is ≥65%.

[0024] The preparation method of the above pickled Chinese cabbage includes the following steps: S1. Salt pickling in the pool: Load the pool in the way of one layer of vegetable embryo and one layer of salt, and seal for salt pickling; S2. Secondary pickling in the jar: After washing the salted vegetable embryo, transfer it into the jar, add brine with a salinity of 8-13% and the bacterial liquid or bacterial sludge made from Lactiplantibacillus plantarum TLP03 for fermentation to obtain pickled Chinese cabbage; among them, rock sugar is added to the brine; the fermentation period is 15-25 d; calculated by the weight of the brine, the inoculation amount of the bacterial liquid is (8-12) g / kg; the inoculation amount of the bacterial sludge is (0.5-1.5) g / kg.

[0025] In a preferred mode, the concentration of the brine is 8-10%, preferably 8%.

[0026] Compared with the prior art, the present invention has the following advantages and beneficial effects: 1. The Lactiplantibacillus plantarum TLP03 provided by the present invention can grow and metabolize in an environment with a pH of 3-5 and high salt (8%-10%).

[0027] 2. Preparing the Lactiplantibacillus plantarum TLP03 provided by the present invention into a bacterial sludge or bacterial liquid and adding it to the sauerkraut fermentation system can increase the contents of lactic acid, acetic acid, glutamic acid and hexanal in sauerkraut. Among them, the highest content of lactic acid can reach 15.8 g / kg, the highest content of acetic acid can reach 1.02 g / kg, the highest content of glutamic acid can reach 0.15 g / kg, and the highest content of hexanal can reach 1.12 μg / kg. Description of the Drawings

[0028] The drawings described herein are used to provide a further understanding of the embodiments of the present invention, form a part of this application, and do not limit the embodiments of the present invention. In the drawings: Figure 1 It is the colony map of Lactiplantibacillus plantarum TLP03 in Example 2 of the present invention; Figure 2 It is the cell morphology map of Lactiplantibacillus plantarum TLP03 under microscope observation in Example 2 of the present invention. Detailed Embodiments

[0029] To make the objectives, technical solutions and advantages of the present invention clearer and more understandable, the present invention will be further described in detail below in conjunction with the embodiments. The illustrative embodiments and descriptions of the present invention are only used to explain the present invention and do not limit the present invention. The following described embodiments are part of the embodiments of the present invention, not all of the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts fall within the scope of protection of the present invention.

[0030] In the following description, a large number of specific details are set forth in order to provide a thorough understanding of the present invention. However, it is obvious to those of ordinary skill in the art that: these specific details do not have to be used to implement the present invention. In other embodiments, well-known structures, materials or methods are not specifically described in order to avoid obscuring the present invention. The materials, instruments and reagents used in the following embodiments can be obtained from commercial sources without special instructions. The technical means used in the embodiments are conventional means well-known to those skilled in the art without special instructions.

[0031] In order to improve the taste of sauerkraut, in this embodiment, the strain - Lactiplantibacillus plantarum TLP03 is obtained through screening. The Lactiplantibacillus plantarum TLP03 can be prepared into a preparation, and the form of the preparation includes a bacterial agent, bacterial powder or bacterial liquid. The Lactiplantibacillus plantarum TLP03 or the prepared preparation can be added to the sauerkraut fermentation system, which can increase the contents of lactic acid, acetic acid, glutamic acid and hexanal in sauerkraut, thereby improving the taste of sauerkraut.

[0032] Preferably, when applying Lactiplantibacillus plantarum TLP03, Lactiplantibacillus plantarum TLP03 is first prepared into a bacterial sludge or bacterial liquid, and then the bacterial sludge or bacterial liquid is inoculated into pickled Chinese sauerkraut soaked in brine for fermentation. Based on the weight of the brine, the inoculation amount of the bacterial liquid is (8 - 12) g / kg; the inoculation amount of the bacterial sludge is (0.5 - 1.5) g / kg. Preferably, based on the weight of the brine, the inoculation amount of the bacterial liquid is (10 - 12) g / kg; the inoculation amount of the bacterial sludge is (1.0 - 1.5) g / kg. Preferably, based on the weight of the brine, the inoculation amount of the bacterial liquid is 10 g / kg; the inoculation amount of the bacterial sludge is 1.0 g / kg.

[0033] A method for preparing pickled Chinese sauerkraut fermented by Lactiplantibacillus plantarum TLP03 comprises the following steps: S1. Salt pickling in the pool: The pool is filled in the way of laying one layer of vegetable embryos and one layer of salt, and then sealed for salt pickling; the vegetable embryos include large-leaf mustard, small-leaf mustard, Chinese cabbage, pakchoi, radish and chili; S2. Secondary pickling in the jar: After washing the salt-pickled vegetable embryos, transfer them into the jar, add brine with a salinity of 8 - 13% and the bacterial liquid or bacterial sludge made from Lactiplantibacillus plantarum TLP03 for fermentation to obtain pickled Chinese sauerkraut; among them, rock sugar is added to the brine; the fermentation period is 15 - 25 d; based on the weight of the brine, the inoculation amount of the bacterial liquid is (8 - 12) g / kg; the inoculation amount of the bacterial sludge is (0.5 - 1.5) g / kg; S3. Packaging: Pack the fermented pickled Chinese sauerkraut in bags and perform sterilization treatment.

[0034] Preferably, the concentration of the brine is 8 - 10%, and further preferably, the concentration of the brine is 8%.

[0035] The pickled Chinese sauerkraut prepared above can be used in the preparation of compound condiments, and the compound condiments include but are not limited to pickled fish seasonings, pickled Chinese sauerkraut hot pot bases and pickled Chinese sauerkraut rice noodle seasonings, etc.

[0036] Example 1: The breeding of the strain comprises the following steps: 1). Primary screening: The applicant obtained a total of 4 samples of the end-stage vegetable samples and water samples of large-leaf mustard after the first salt pickling from a food factory in Mianyang (Sichuan Jijia Weilai Food Co., Ltd.) on May 28, 2024. Take 25 g of the sample and add it to 225 mL of sterile physiological saline, shake and elute for more than 1 min, take 1 mL of the eluate and add it to 9 mL of sterile physiological saline for 10-fold serial dilution to obtain dilution solutions with dilution factors of 10 -2 ; Select the undiluted solution, 10 -1 , 10 -2For three gradients, take 100 µL of the gradient-diluted solution and evenly spread it on the MRS medium plate. Invert the plate after spreading and place it in an anaerobic chamber. At the same time, place an anaerobic gas-generating bag equivalent to the volume of the anaerobic chamber. After sealing the anaerobic chamber, place it in an incubator at 35 - 39 °C and incubate for 24 - 72 h. Take out the plate when single colonies can be observed. Select single colonies with typical lactic acid bacteria characteristics based on the colony morphology on the MRS medium plate. Use a sterile inoculation loop to pick one loop of each marked single colony and streak it on the MRS medium plate. After purification, determine the cell morphology of the strain by Gram staining microscopy. Finally, obtain 30 suspected lactic acid bacteria strains; use a sterile inoculation loop to pick single colonies on the purified plate, inoculate them into MRS broth medium, and incubate in a constant-temperature incubator at 37 °C for 24 - 48 h. After vortexing and mixing the test tube containing the bacterial suspension, take the bacterial suspension and mix it with 40% glycerol in a 1:1 ratio in a centrifuge tube. Mix well, wrap the centrifuge tube mouth with sealing film, and place it in a -20 °C refrigerator for preservation.

[0037] Among them, the composition of the MRS agar medium is: peptone 10.0 g / L, beef extract powder 10.0 g / L, glucose 20.0 g / L, yeast extract powder 5.0 g / L, sodium acetate 3.02 g / L, dipotassium hydrogen phosphate 1.16 g / L, magnesium sulfate 0.05 g / L, ammonium citrate 2.0 g / L, manganese sulfate 0.03 g / L, Tween 80 1 mL / L, agar powder 15 g / L.

[0038] Among them, the composition of the MRS broth medium is: peptone 10.0 g / L, beef powder 5.0 g / L, glucose 20.0 g / L, yeast powder 4.0 g / L, sodium acetate 5.0 g / L, dipotassium hydrogen phosphate 2.0 g / L, magnesium sulfate 0.2 g / L, ammonium citrate 2.0 g / L, manganese sulfate 0.05 g / L, Tween 80 1 mL / L.

[0039] 2) Determination of the tolerance ability of the strain: Inoculate the strain to be tested in the preservation tube into MRS broth medium and activate it in a constant-temperature incubator at 37 °C for 24 h, activating two generations each time to ensure the viability of the strain.

[0040] Determination of the acid tolerance ability of the strain: Use 0.1 mol / L HCl solution to adjust the pH of the prepared MRS broth medium to 3, 4, 5, and 6, autoclave at 121 °C for 15 min, and after cooling, inoculate the activated strain into the above MRS broth medium at an inoculation amount of 2% by volume fraction and place it in a constant-temperature incubator at 37 °C for 24 h. Use the MRS broth medium without inoculation as a blank control. Measure the absorbance values of the culture solution at 600 nm at 0 h and 24 h of incubation, and calculate lg(OD 24h / OD 0h), determine the growth rate of the strain in different acidic environments.

[0041] Determination of the salt tolerance of the strain: Sterilize MRS broth media containing 6%, 8%, and 10% sodium chloride at 121 °C for 15 min. After cooling, inoculate the activated strain into the above MRS broth media at an inoculation volume fraction of 2%, and place them in a constant temperature incubator at 37 °C for 24 h. Use the MRS broth media without inoculation as a blank control. Measure the absorbance values of the culture broth at 600 nm at 0 h and 24 h of culture, and calculate lg(OD 24h / OD 0h ), and determine the growth rate of the strain in environments with different salt contents.

[0042] Determination of the low-temperature tolerance of the strain: Sterilize MRS broth media at 121 °C for 15 min. After cooling, inoculate the activated strain into the above MRS broth media at an inoculation volume fraction of 2%, and place them in a constant temperature incubator at 10 °C for 24 h. Use the MRS broth media without inoculation as a blank control. Measure the absorbance values of the culture broth at 600 nm at 0 h and 24 h of culture, and calculate lg(OD 24h / OD 0h ), and determine the growth rate of the strain in a low-temperature environment.

[0043] Test results: Some strains can grow and metabolize in environments with pH 3 - 5, high salt (8% - 10%), and low temperature (10 °C). Based on the comprehensive test results of various tolerance abilities, 5 strains with relatively strong comprehensive tolerance abilities were finally selected.

[0044] 3), Determination of the acid production ability of the strain: Number the 5 selected strains with relatively strong tolerance as 1, 2, 3, 4, and 5, and activate them in the same way as in step 2); inoculate the activated strains into MRS broth media at an inoculation volume fraction of 2%, and place them in a constant temperature incubator at 37 °C for 24 h. Evaluate their acid production ability by the pH method and determine their organic acid content by high-performance liquid chromatography. The obtained results are shown in Table 1. The organic acid determination method is carried out according to "GB 5009.157 - 2016 National Food Safety Standard Determination of Organic Acids in Foods". Since lactic acid bacteria mainly produce lactic acid and acetic acid, directly detect the ability to produce lactic acid and acetic acid. Use the strain with the highest production of lactic acid and acetic acid (number 3) for subsequent research, and name the strain TLP03.

[0045] Table 1 Determination of the acid production ability of the strain

[0046] It can be seen from the data in Table 1 that the strains numbered 1-5 all have strong acid-producing ability, and the strain numbered 3 has the best acid-producing ability.

[0047] Example 2: Identification of strain TLP03 1), Morphological identification: Inoculate the strain TLP03 screened in Example 1 on MRS agar medium, place it in a 37 °C incubator and streak culture for 24 h to form white, round or near-round, small colonies with smooth edges; observe under a 100-fold oil microscope, the strain morphology is short rod-shaped, arranged neatly, with rounded ends at both ends and no spores. As Figure 1 、 Figure 2 shown.

[0048] 2), Physiological and biochemical identification Inoculate the strain TLP03 in the preservation tube on MRS agar medium, place it in a 37 °C incubator and streak culture for 24 h. Prepare lactic acid bacteria biochemical identification tubes (esculin biochemical tube, cellobiose biochemical tube, lactose biochemical tube, sucrose biochemical tube, raffinose biochemical tube, mannitol biochemical tube, maltose biochemical tube, sorbitol biochemical tube, inulin biochemical tube, salicin biochemical tube, sodium hippurate biochemical tube). Use a glass slide to cut open the upper end of the biochemical tube, and use a sterile toothpick to pick up single colonies on the plate and inoculate them into each biochemical tube respectively. After the bacteria attached to the toothpick completely fall into the biochemical solution, block the upper end of the biochemical tube with a cotton strip, and invert the culture according to the culture temperature and time requirements of each biochemical tube. After the culture is completed, record the color and state of the biochemical tube.

[0049] The results are shown in Table 2: Table 2

[0050] 3) Molecular biology identification Using the total DNA of strain TLP03 as a template, PCR amplification was carried out with the 16s rDNA universal primers 27F (5’-AGAGTTTGATCCTGGCTCAG-3’) and 1492R (5’-CTACGGCTAC CTTGTTACGA-3’). Entrusted Shanghai Huace to amplify and sequence the 16S rDNA of this bacterium. After obtaining the 16S rDNA sequence of this strain (shown in SEQ ID NO.1), use BLAST on the NCBI website to retrieve the 16S rDNA gene sequences of related strains in GenBank and conduct homology comparison, and the homology is 99%. Combining the biochemical identification results, it is considered that strain TLP03 is Lactiplantibacillus plantarum ( Lactiplantibacillus plantarum ), and it is tentatively named Lactiplantibacillus plantarum ( Lactiplantibacillus plantarum)(TLP03), deposited with the China Center for Type Culture Collection, with the deposit name of TLP03 and the deposit number of CCTCC NO: M20242276; Deposit date: October 21, 2024, Deposit address: Wuhan University, Wuhan, China, Zip code: 430072.

[0051] SEQ ID NO.1:

[0052] Example 3: Preparation of the bacterial sludge and bacterial liquid containing Lactobacillus plantarum TLP03: Inoculate the Lactobacillus plantarum TLP03 into MRS broth medium and activate it at 36 °C for 25 h to obtain a seed liquid; inoculate the seed liquid into another MRS broth medium at an inoculation amount of 2% (v / v) and activate it at 36 °C for 25 h to obtain a fermentation liquid, i.e., the bacterial liquid.

[0053] Centrifuge the bacterial liquid at a centrifugation temperature of 5 °C, a centrifugation speed of 8000 rpm, and a centrifugation time of 8 min, and collect the lower-layer bacterial cell precipitate after centrifugation, i.e., the bacterial sludge is obtained.

[0054] Example 4: Effect of different pH values on the growth and acid production of Lactobacillus plantarum TLP03 Prepare MRS broth medium, adjust the pH with 0.1 mol / L HCl solution, and statically culture it at 37 °C for a certain period of time. If the bottom of the Erlenmeyer flask is covered with bacteria, the strain can grow under this pH condition. After culturing under the same conditions for 24 h, mix the bacterial liquid, and measure the OD of the bacterial liquid at 0 h and 24 h 600 value and the titratable acid content of the supernatant to characterize the growth and acid production of the strain under acidic conditions. The results are shown in Table 3. In the culture media with different pH values, the OD 600nm values all increased to varying degrees, indicating that the strain has low pH tolerance and is suitable for growth and metabolism in the environment of sauerkraut fermentation process.

[0055] Table 3

[0056] Example 5: Effect of different salt contents on the growth and acid production of Lactobacillus plantarum TLP03 Prepare MRS broth medium and add sodium chloride to adjust the salt concentration, and statically culture it at 37 °C for a certain period of time. If the bottom of the Erlenmeyer flask is covered with bacteria, the strain can grow under this salt concentration condition. After culturing under the same conditions for 24 h, mix the bacterial liquid, and measure the OD of the bacterial liquid at 0 h and 24 h 600 value and the titratable acid content of the supernatant to characterize the growth and acid production of the strain under acidic conditions. The results are shown in Table 4: The strain has high salt concentration tolerance and is suitable for growth and metabolism in the environment of sauerkraut fermentation process.

[0057] Table 4

[0058] Note: The NaCl concentration in Table 4 is the mass-volume ratio. 6% NaCl means 6 g of sodium chloride in a 100 mL system.

[0059] Example 6: Application of Lactiplantibacillus plantarum TLP03 in the process of pickled Chinese cabbage fermentation: A method for preparing pickled Chinese cabbage, comprising the following steps: S1. Salt pickling in the pool: For every 100 kg of fresh large-leaf mustard greens, use 15 kg of salt; during the process of loading into the pool, layer the vegetable embryos and salt, and evenly sprinkle the salt according to the weight of the fresh vegetables. After the vegetable embryos are salted and produce brine, perform at least one brine circulation every day. After filling the salt pickling pool, sprinkle a certain amount of salt on the surface, cover it with a thick transparent plastic film, and then evenly spread river sand on it and seal it tightly.

[0060] S2. Secondary pickling in the altar: Transfer the vegetable embryos out of the pool to the processing point, remove abnormal vegetables such as those that turn black or have ventilation problems during the salt pickling process, use a knife to cut off the vegetable heads, old stems, and yellow leaves, and select foreign matters. After processing, evenly put them into a cleaning machine, and wash out foreign matters such as sediment, dust, and leaves mixed in the leaf stalks and vegetable bodies through the cleaning machine until the brine exuded by the vegetable bodies is clear and not turbid; evenly transfer the washed salted vegetable embryos into the altar for fermentation. The total loading capacity of the pickling altar is 300 kg, and each altar contains 150 kg of vegetable embryos. After weighing the vegetable embryos and putting them into the altar, directly add brine to the edge of the altar; the fermentation period is 15 days.

[0061] Control group 1: Ferment with brine with a salinity of 8%, and rock sugar is added to the brine. The addition amount of rock sugar is 0.67% of the weight of the vegetable embryos.

[0062] Control group 2: Ferment with brine with a salinity of 8% and a bacterial liquid made from Lactiplantibacillus plantarum CICC 20242. Rock sugar is added to the brine. The addition amount of rock sugar is 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial liquid is 10 g / kg.

[0063] Experimental group 1: Ferment with brine with a salinity of 8% and a bacterial liquid made from Lactiplantibacillus plantarum TLP03. Rock sugar is added to the brine. The addition amount of rock sugar is 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial liquid is 10 g / kg.

[0064] Experimental group 2: Ferment with brine with a salinity of 8% and a bacterial mud made from Lactiplantibacillus plantarum TLP03. Rock sugar is added to the brine. The addition amount of rock sugar is 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial mud is 1.0 g / kg.

[0065] Experimental group 3: Fermentation was carried out by adding brine with a salinity of 8% and a bacterial solution made from Lactiplantibacillus plantarum TLP03. Rock sugar was added to the brine, and the addition amount of rock sugar was 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial solution was 8 g / kg.

[0066] Experimental group 4: Fermentation was carried out by adding brine with a salinity of 10% and a bacterial solution made from Lactiplantibacillus plantarum TLP03. Rock sugar was added to the brine, and the addition amount of rock sugar was 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial solution was 8 g / kg.

[0067] Experimental group 5: Fermentation was carried out by adding brine with a salinity of 10% and a bacterial solution made from Lactiplantibacillus plantarum TLP03. Rock sugar was added to the brine, and the addition amount of rock sugar was 0.67% of the weight of the vegetable embryos; based on the weight of the brine, the inoculation amount of the bacterial solution was 10 g / kg.

[0068] Refer to "GB 5009.157-2016 National Food Safety Standard - Determination of Organic Acids in Foods" to determine the contents of lactic acid and acetic acid in the fermentation system, and refer to "GB 5009.124-2016 National Food Safety Standard - Determination of Amino Acids in Foods" and "GB / T 11538-2006 General Method for Capillary Column Gas Chromatographic Analysis of Essential Oils" to determine the contents of glutamic acid and hexanal in the fermentation system. The results are shown in Table 5: Table 5

[0069] It can be seen from the data in Table 5 that: 1) The addition amount of the bacterial solution and the brine concentration both have an impact on the content of the fermentation products; 2) Compared with the existing Lactiplantibacillus plantarum CICC 20242, the strains screened in this example have greatly improved the ability to produce acetic acid, glutamic acid and hexanal when used for sauerkraut fermentation.

[0070] The specific embodiments described above have further detailed the purpose, technical solution and beneficial effects of the present invention. It should be understood that the above description is only the specific embodiments of the present invention and is not used to limit the protection scope of the present invention. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A plant lactobacillus TLP03, characterized in that Lactobacillus plantarum ( Lactiplantibacillus plantarum ) TLP03 is deposited in China Center for Type Culture Collection, its deposit name is TLP03, and its deposit number is CCTCCNO: M20242276.

2. A preparation containing Lactobacillus plantarum TLP03 as claimed in claim 1, characterized in that: The preparations include bacterial agents, bacterial mud, bacterial powder or bacterial liquid.

3. Application of plant lactobacillus TLP03 as claimed in claim 1 in a sauerkraut fermentation process.

4. The use according to claim 3, characterized in that: Firstly, the Lactobacillus plantarum TLP03 is prepared into bacterial mud or bacterial liquid, and then the bacterial mud or bacterial liquid is inoculated into sauerkraut soaked in salt water for fermentation.

5. The use according to claim 4, characterized in that: Based on the weight of the salt water, the inoculation amount of the bacterial liquid is (8-12) g / kg; the inoculation amount of the bacterial mud is (0.5-1.5) g / kg.

6. The use according to claim 4, characterized in that: The preparation process of the bacterial solution is as follows: Inoculating the plant lactobacillus into an MRS broth medium, activating the medium at 35-39° C. for 20-26 hours, and obtaining a seed solution; Inoculate the seed liquid in another MRS broth medium at an inoculation rate of 1-5%, activate at 35-39° C. for 20-26 h to obtain a fermentation liquid, i.e., the bacterial liquid; The preparation process of the bacterial mud is as follows: The bacterial liquid is centrifuged at a centrifugal temperature of 2-6° C., a centrifugal speed of 6000-10000 rpm, and a centrifugal time of 6-10 min. After centrifugation, the lower bacterial precipitate is collected to obtain the bacterial mud.

7. Use of the preparation as claimed in claim 2 in a sauerkraut fermentation process.

8. A product, characterized in that The product comprises sauerkraut or a composite condiment containing the sauerkraut; The sauerkraut is prepared by fermentation of the Lactobacillus plantarum TLP03 according to claim 1 or the preparation according to claim 2; The composite seasoning comprises pickled fish seasoning, pickled vegetable hot pot seasoning or pickled vegetable rice noodle seasoning.

9. The product according to claim 8, characterized in that The preparation process of the sauerkraut comprises the following steps: S1. Salting in the pool: The pool is filled with a layer of vegetable embryos and a layer of salt, and then sealed for salting; S2, secondary soaking in a jar: washing the salted vegetable embryos and transferring them into a jar, adding salt water with a salinity of 8-13% and a bacterial liquid or bacterial mud made from the Lactobacillus plantarum TLP03 for fermentation to obtain sauerkraut; wherein crystal sugar is added to the salt water; and the fermentation period is 15-25 days.

10. The product according to claim 9, characterized in that Based on the weight of the salt water, the inoculation amount of the bacterial liquid is (8-12) g / kg; the inoculation amount of the bacterial mud is (0.5-1.5) g / kg.

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