Multifunctional plant lactobacillus and application thereof in sour leaf mustard processing

Abstract

The invention discloses a multifunctional plant lactobacillus and an application of the multifunctional plant lactobacillus in sour mustard processing, the name of the multifunctional plant lactobacillus is plant lactobacillus LJ036, and the preservation number of the multifunctional plant lactobacillus in the China Center for Type Culture Collection (CCTCC) is CCTCC NO: M 20241710. The multifunctional plant lactobacillus can be applied to sour leaf mustard processing. According to the plant lactobacillus LJ036, the nitrite content in the leaf mustard fermentation process can be obviously reduced, the nitrite content in the fermentation process is lower than 1.3 mg / kg, the acetoin content of the sour leaf mustard can be increased by the plant lactobacillus LJ036 and is 4.13 times that of natural fermentation, the fermentation time can be obviously shortened by the plant lactobacillus LJ036, and the plant lactobacillus LJ036 has a good application prospect.

Description

Technical Field

[0001] This invention belongs to the field of food processing technology and relates to multifunctional Lactobacillus plantarum and its application in the processing of pickled mustard greens. Specifically, it relates to a strain of Lactobacillus plantarum LJ036 that degrades nitrite and produces aroma and its application in the processing of pickled mustard greens. Background Technology

[0002] Pickled vegetables have a history of over 3,000 years in China and are a traditional fermented vegetable food. Rich in organic acids, vitamins, dietary fiber, and other nutrients, they are typically served as a side dish or appetizer. Pickled mustard greens, a traditional fermented food in China with a history spanning thousands of years, are widely loved for their sour, fragrant, crisp, and appetizing taste. The fermentation process of pickled mustard greens involves a variety of aroma-producing microorganisms. Furthermore, the fermentation of wine, yogurt, soy sauce, and vinegar also involves abundant aroma-producing microorganisms, giving the products various flavors.

[0003] Nitrites are produced during the pickling process of mustard greens. The nitrite content varies depending on the raw vegetables, the salt content and temperature of the pickling process, and the types of nitrate-reducing bacteria. As the pickling time increases, the nitrite content first gradually rises to a peak, then gradually decreases. If the curing time is insufficient or there are many other bacteria during pickling, nitrite levels can easily exceed safe limits. Therefore, food safety issues related to pickled mustard greens cannot be ignored. Nitrite content is a crucial indicator that cannot be overlooked during the processing and storage of pickled mustard greens. Excessive nitrite intake can cause various health problems. Consuming a single dose can lead to acute toxicity such as hypoxemia. Furthermore, nitrites combine with amines in the body to form nitrosamines, which are potent carcinogens and increase the risk of cancer. Therefore, controlling nitrite production in pickled mustard greens is particularly important. In 2017, the International Agency for Research on Cancer (IARC) of the World Health Organization classified nitrates or nitrites ingested under endogenous nitrosation conditions as Group 2A carcinogens. According to my country's regulations on the limits of contaminants in food, the limit for nitrite in pickled vegetables and their products shall not exceed 20 mg / kg.

[0004] Lactic acid bacteria are Gram-positive bacteria, generally considered safe. They ferment sugars to produce acid and play a dominant role in the fermentation process of pickled mustard greens. Studies have shown that some lactic acid bacteria screened from fermented vegetables can reduce the nitrite content in pickled vegetables, while others have aroma-producing effects. Furthermore, lactic acid bacteria also have probiotic effects such as preventing diarrhea and peptic ulcers, lowering serum cholesterol, alleviating lactose intolerance, and improving immunity. Zhu Junli et al. isolated a strain of *Lactobacillus plantarum* from homemade fermented pickles, which achieved a 95.6% degradation rate of 125 mg / L nitrite after 72 hours of cultivation. Wang Ying et al. isolated a strain of *Lactobacillus plantarum* from homemade fermented pickled cowpeas, which achieved a 91.64% degradation rate of 200 mg / L nitrite after 48 hours of cultivation. Lü Xinran screened high-AI-2 signaling bacteria strains from 16 lactic acid bacteria derived from traditional fermented foods and carp intestines and analyzed their aroma-producing capabilities. One high-AI-2 producing strain, DBM2-4, was identified as *Lactobacillus plantarum* through physiological and biochemical analysis and 16S rRNA testing. Compared to the low-AI-2 producing strain YF-7, strain DBM2-4, after being cultured at 37℃ for 72 hours, showed increased yields of the flavor compounds dimethylglyoxal (DME) and acetaldehyde by 3.42 mg / L and 5.86 mg / L, respectively. However, currently, there are very few multifunctional superior strains developed. Screening for multifunctional strains is beneficial for simultaneously improving multiple properties of fermented vegetables (such as safety and product flavor quality), and represents a research and development trend in fermented vegetable production. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to overcome the shortcomings of the prior art and provide a multifunctional plant lactobacillus that can degrade nitrite and produce aroma while fermenting mustard greens, and its application in the processing of sour mustard greens.

[0006] To solve the above-mentioned technical problems, the present invention adopts the following technical solution.

[0007] A multifunctional plant lactobacillus strain, named *Lactiplantibacillus plantarum* LJ036, has the accession number CCTCCNO: M 20241710 at the China Center for Type Culture Collection (CCTCC), located at 299 Bayi Road, Wuchang District, Wuhan, Hubei Province, China (within the campus of Wuhan University). The accession date is July 31, 2024.

[0008] The colony characteristics of *Lactiplantibacillus plantarum* LJ036 of this invention are as follows: colonies are raised, milky white, round, medium-sized, moist, with neat, opaque edges, and a diameter of 3 mm to 3.5 mm. It is Gram-positive and catalase-negative. The 16S rDNA sequence of this *Lactiplantibacillus plantarum* LJ036 is shown in SEQ ID NO.1. Through 16S rDNA sequence alignment analysis, it was found that the strain of this invention shares 100% homology with *Lactiplantibacillus plantarum*. Based on the bacterial morphology, growth characteristics, and physiological and biochemical identification results, this lactic acid bacterium LJ036 is confirmed to be *Lactiplantibacillus plantarum*.

[0009] As a general technical concept, the present invention also provides an application of the above-mentioned multifunctional plant lactobacillus in the processing of sour mustard greens.

[0010] Preferably, the above application includes the following steps:

[0011] S1. Raw material cleaning: After the mustard greens are cleaned and pesticide residues are removed, they are dried or shaken to remove surface water.

[0012] S2. Raw material sterilization treatment: The mustard greens obtained in step S1 are sterilized by pulse ultrasound.

[0013] S3. Activation of multifunctional plant lactobacillus: The multifunctional plant lactobacillus is activated to obtain a live bacterial solution;

[0014] S4. Mustard green pickling and inoculation: Weigh the mustard greens, pickle them with 6% to 8% salt by weight of the mustard greens, pack them into a fermentation container and press them down layer by layer, with a filling volume of 70% to 80%. Inoculate the live bacteria solution obtained in step S3 into the pickled mustard greens, and then add salt water with a salt content of 6wt% to 8wt% until the mustard greens are submerged.

[0015] S5. Fermentation: Control the temperature at 25℃~30℃, and ferment for 12d~16d;

[0016] S6. Termination of fermentation: When the total acidity of the mustard greens reaches 8.0g / kg~10.0g / kg, the greens are harvested to obtain sour mustard greens;

[0017] S7. Preservation of pickled mustard greens: The pickled mustard greens obtained in step S6 are preserved using a fence-like anti-corrosion and preservation technology to complete the processing of pickled mustard greens.

[0018] In the above application, preferably, the activation process in step S3 is as follows: The multifunctional *Lactobacillus plantarum* is first cultured on an MRS solid medium slant, and then cultured on an MRS liquid medium at 36℃~38℃ for 24h~48h. The resulting fermentation broth is centrifuged at 3℃~5℃ and 5000rpm~7000rpm for 10min~15min, and the bacterial cells are collected. The cells are washed 2~3 times with physiological saline, and then resuspended in physiological saline to adjust the bacterial concentration to 10. 7 cfu / mL ~10 8 The cfu / mL concentration was used to obtain a live bacterial culture, which was then refrigerated at 4℃~6℃.

[0019] In the above application, preferably, in step S4, the pickling time is 2h to 3h, and the volume of the inoculated live bacteria liquid / (the total volume of the live bacteria liquid + the pickled mustard greens + the brine) = 1% to 2%.

[0020] In the above application, preferably, in step S7, the preservation process of the fence anti-corrosion and preservation technology is as follows: 0.5g / kg to 1.0g / kg of nisin is added to the sour mustard greens, vacuum quantitatively packaged, sterilized under a pressure of 500MPa to 600MPa for 5min to 7min, and cold-chain stored and transported at 4℃ to 6℃; the shelf life is more than 6 months.

[0021] In the above application, preferably, in step S1, the process of removing pesticide residues is as follows: the washed mustard greens are treated with high-concentration ozone water, wherein the ozone concentration of the high-concentration ozone water is 5mg / L to 10mg / L, and the treatment time is 10min to 20min.

[0022] In the above application, preferably, in step S2, the pulsed ultrasound is a single-frequency pulsed ultrasound or a dual-frequency pulsed ultrasound. The frequency of the single-frequency pulsed ultrasound is 80 kHz to 100 kHz, and the frequency of the dual-frequency pulsed ultrasound is 80 kHz to 100 kHz. Each ultrasound treatment lasts 20 to 30 seconds, with an interval of 5 to 6 seconds, for a total duration of 20 to 30 minutes.

[0023] In the above application, preferably, in step S4, the preparation process of the brine is as follows: prepare 1 to 1.5 times the weight of the mustard greens in boiled and cooled water, add salt, and prepare brine.

[0024] Compared with the prior art, the advantages of the present invention are as follows:

[0025] 1. Based on screening a large number of lactic acid bacteria resources, this invention uses the nitrite degradation rate and acetoin content in fermented mustard juice as evaluation indicators, combined with acid production capacity, to obtain a *Lactobacillus plantarum* strain LJ036 that exhibits excellent nitrite degradation and aroma production performance. This *Lactobacillus plantarum* LJ036 can be used to process and produce sour mustard greens, possessing both excellent nitrite degradation capacity and aroma production effect. Lactobacillus plantarum LJ036 significantly reduced nitrite content during mustard fermentation, achieving a nitrite degradation rate of up to 96.2%. Nitrite levels remained below 1.3 mg / kg throughout the fermentation process. The acetoin content in the fermented mustard juice inoculated with LJ036 reached 43.3 ± 0.16 mg / L, and the pH decreased to 3.51. The acetoin content after 14 days of fermentation with LJ036 was 4.13 times that of natural fermentation (CK) after 24 days, effectively improving the aroma and quality of sour mustard while shortening the fermentation time by 41.7% compared to natural fermentation.

[0026] This invention collected different types of fermented vegetables from various regions of Hunan Province. 295 single bacterial strains were isolated and screened from samples such as homemade pickled vegetables, pickled mustard greens, and chopped chili peppers from Hunan households. Gram staining and catalase experiments preliminarily identified 85 lactic acid bacteria strains. Screening was conducted based on the bacteria's ability to degrade nitrite in MRS medium, the acetoin content in fermented mustard green juice, and its acid-producing capacity. The resulting multifunctional *Lactobacillus plantarum* LJ036 was obtained and applied to the processing of pickled mustard greens, providing technical support for meeting the demand for superior bacterial strains that improve the safety and quality of pickled mustard green production.

[0027] 2. To overcome the shortcomings of existing heat sterilization technologies for pickled mustard greens, which affect the flavor and heat-sensitive components of the product, this invention employs non-heat sterilization technology during the processing of pickled mustard greens. It constructs a four-dimensional barrier preservation technology combining "high acid + ultra-high pressure + preservative + low temperature." Under this four-dimensional barrier preservation technology—containing a total acid of 8.0g / kg to 10.0g / kg, ultra-high pressure of 500MPa to 600MPa for 5 to 7 minutes, nisin of 0.5g / kg to 1.0g / kg, and a low temperature of 4℃ to 6℃—the product exhibits a significant synergistic antibacterial and enhancing effect compared to single-dimensional, two-dimensional, and three-dimensional methods. This not only maintains the tender, crisp, and tangy taste and rich fermented aroma of the pickled mustard greens but also extends its shelf life to over 6 months.

[0028] Preservation of biological materials

[0029] A multifunctional plant lactobacillus strain, named *Lactiplantibacillus plantarum* LJ036, has the accession number CCTCCNO: M 20241710 at the China Center for Type Culture Collection (CCTCC), located at 299 Bayi Road, Wuchang District, Wuhan, Hubei Province, China (within the campus of Wuhan University). The accession date is July 31, 2024. Attached Figure Description

[0030] Figure 1 The images show the colony morphology (left) and Gram staining (right) of *Lactobacillus plantarum* LJ036 in Example 1 of this invention.

[0031] Figure 2 This is the phylogenetic tree of Lactobacillus plantarum LJ036 in Example 1 of the present invention.

[0032] Figure 3 This is a comparative graph of the effects of the naphthylethylenediamine hydrochloride method on *Lactobacillus plantarum* LJ036, *Lactobacillus plantarum* LJ065, *Lactobacillus plantarum* JC080, and the control group CK in Example 1 of the present invention. In this graph, tubes 1 and 2 contain *Lactobacillus plantarum* LJ036, tubes 3 and 4 contain *Lactobacillus plantarum* LJ065, tubes 5 and 6 contain *Lactobacillus plantarum* JC080, and tubes 7 and 8 contain the control group CK.

[0033] Figure 4 This is a comparison chart of the content of acetoin in fermented mustard juice by *Lactobacillus plantarum* LJ036 and other plant lactic acid bacteria LJ065, JC080, JC091, JC045, and JC052 in Example 1 of the present invention. Detailed Implementation

[0034] The present invention will be further described below with reference to the accompanying drawings and specific preferred embodiments, but this does not limit the scope of protection of the present invention. All materials and instruments used in the following embodiments are commercially available.

[0035] Example 1

[0036] The present invention discloses a multifunctional plant lactobacillus strain, named *Lactiplantibacillus plantarum* LJ036, with accession number CCTCCNO: M 20241710 at the China Center for Type Culture Collection (CCTCC), located at 299 Bayi Road, Wuchang District, Wuhan, Hubei Province, China (within the campus of Wuhan University). The accession date is July 31, 2024.

[0037] The isolation, purification, and identification of Lactobacillus plantarum LJ036 in this embodiment:

[0038] Weigh 25g of each sample, add 225mL of sterile physiological saline, homogenize, and then perform a 10-fold serial dilution. Take 10... -5 10 -6 0.1 mL of each dilution was plated onto MRS solid medium (containing 0.2% glucose, 0.4% yeast extract, 1% peptone, 1.5% agar powder, and 0.5% beef meal) containing calcium carbonate and incubated at 37°C for 48 h. Typical single colonies with large calcium dissolution zones were selected and repeatedly streaked onto MRS plates to obtain pure strains. Gram staining and catalase tests were performed on the strains. Colonies with distinct calcium dissolution zones, Gram-positive staining, and negative catalase reaction (milky white or yellowish-white with a smooth surface) were preliminarily identified as lactic acid bacteria. The strains were stored at -80°C in MRS medium supplemented with 20% glycerol.

[0039] The above-mentioned strains underwent morphological observation, physiological and biochemical experiments, and 16S rDNA amplification and sequencing. All identification results were comprehensively analyzed. Specific identification results are as follows:

[0040] Morphological observation results: Colonies were raised, milky white, round, medium-sized, moist, with neat, opaque edges, and a diameter of 3mm–3.5mm. Gram-positive, catalase-negative. Colony morphology and Gram-stained photographs of strain LJ036 are shown below. Figure 1 .

[0041] The physiological and biochemical characteristics of strain LJ036 are shown in Table 1. The strain's ability to utilize 50 carbon sources was determined using API test strips, and strain LJ036 could utilize 22 of these carbohydrates. Finally, strain LJ036 was identified as *Lactiplantibacillus plantarum* using the API LABPlus automated interpretation system.

[0042] Table 1. Physiological and biochemical identification results of Lactobacillus plantarum LJ036

[0043]

[0044]

[0045] 16S rDNA amplification and sequencing analysis: Identification and sequencing were performed by Shanghai Sangon Biotech Co., Ltd.; the sequencing results were compared with existing sequences in the NCBI database, confirming it as *Lactobacillus plantarum*. The sequence obtained by the company is SEQ ID NO.1.

[0046] The genome of *Lactobacillus plantarum* LJ036 was extracted and its 16S rDNA was amplified and sequenced. Sequencing primers were: 27F: AGTTTGATCMTGGCTCAG; 1492R: GGTTACCTTGTTACGACTT. The PCR reaction system (total 25 μL) consisted of: 12.5 μL of 10X PCR Buffer, dNTPs (10 mM each), Taq Plus DNA Polymerase (5 U / μL), and 50 mM MgSO4; 1 μL of primer F (10 μM); 1 μL of primer R (10 μM); 1 μL of template (DNA); and 9.5 μL of ddH2O. The PCR reaction system consisted of: 95℃ pre-denaturation for 5 min, 94℃ denaturation for 30 s, 57℃ annealing for 30 s, and 72℃ extension for 90 s, for a total of 30 cycles, followed by a final extension at 72℃ for 10 min. After purification, the PCR amplification products were sent to a sequencing company for sequencing. The sequencing results (SEQ ID NO.1) were compared with the GeneBank database in NCBI using BLAST. A phylogenetic tree was constructed using MEGA 7.0, and the results are as follows. Figure 2 As shown, the nucleotide sequence of Lactobacillus plantarum LJ036 has 100% homology with the Lactobacillus genus type strain NR113338.1, which has the highest sequence homology in the database.

[0047] Nitrite degradation test results of Lactobacillus plantarum LJ036

[0048] Lactobacillus plantarum LJ036 was inoculated into MRS liquid medium and activated at 37℃ for 24 h. MRS liquid medium was then prepared, pH adjusted to 6.8–7.0, and sodium nitrite was added to prepare a liquid medium containing 200 mg / L NaNO2. Activated Lactobacillus plantarum LJ036, LJ065, and JC080 were inoculated at a 2% inoculation rate into 10 mL of MRS liquid medium containing 200 mg / L NaNO2 and incubated at 37℃ for 18 h. The MRS liquid medium containing 200 mg / L NaNO2 served as the control group (CK). The nitrite degradation rate was calculated as follows: nitrite degradation rate (%) = (nitrite content in the control group - nitrite content in the treatment group) / nitrite content in the control group. The nitrite degradation capacity of Lactobacillus plantarum LJ036 was as high as 96.2%. The degradation effect of Lactobacillus plantarum LJ036 using the naphthylethylenediamine hydrochloride method for nitrite degradation is shown in [reference needed]. Figure 3 .

[0049] Results of the test on the acetoin content in mustard fermentation juice inoculated with Lactobacillus plantarum LJ036

[0050] Weigh a certain amount of mustard greens, add 0.5 times the amount of purified water to make a pulp, filter, and then add 4% glucose, 2% yeast powder, 0.2% dipotassium hydrogen phosphate, and 3% sodium chloride according to the weight of the mustard juice to prepare a mustard juice culture medium. Dispense the medium into 20mL test tubes, autoclave, and set aside. Activate *Lactobacillus plantarum* LJ036 and other screened *Lactobacillus plantarum* strains LJ065, JC080, JC091, JC045, and JC052 in MRS medium, and inoculate them at a ratio of 2% (v / v) into the autoclaved mustard juice culture medium. Incubate at 37℃ for 5 days. Determine the acetoin content in the mustard fermentation juice using the creatine colorimetric method. The results are as follows: Figure 4 As shown, the acetoin content in mustard fermentation juice inoculated with L. plant lactobacillus LJ036 of the present invention is as high as 43.3±0.16mg / L, which is superior to other L. plant lactobacillus strains, and the pH is reduced to 3.51.

[0051] Example 2

[0052] An application of the multifunctional *Lactobacillus plantarum* of the present invention in the processing of pickled mustard greens, specifically *Lactobacillus plantarum* LJ036, and the application method includes the following steps:

[0053] S1. Raw material cleaning: After cleaning the mustard greens, treat them with ozone water with a concentration of 5mg / L for 15 minutes to remove pesticide residues, and then air dry to remove surface water.

[0054] S2. Raw material sterilization treatment: The mustard greens obtained above are placed in a dual-frequency pulse ultrasound device. Each ultrasound treatment lasts for 20 seconds, with a 5-second interval, for a total of 20 minutes. The ultrasound frequency is 100 kHz for sterilization treatment.

[0055] S3. Activation of *Lactobacillus plantarum* LJ036: *Lactobacillus plantarum* LJ036 was first cultured on an MRS solid-state agar slant at 37℃ for 24 h (activation once), and then cultured on MRS liquid medium at 37℃ for 24 h (activation once). The resulting fermentation broth was centrifuged at 4℃ and 5000 rpm for 15 min, and the cells were collected. The cells were washed twice with physiological saline, and then resuspended in physiological saline. The concentration of the broth was adjusted to a viable count of 10-1. 8 The cfu / mL concentration was used to obtain a live bacterial culture, which was then refrigerated at 4°C.

[0056] S4. Mustard green pickling and inoculation: Weigh the mustard greens, pickle them with 8% salt by weight for 3 hours, pack them into a fermentation container, and press them down layer by layer to fill the container to 80%. Inoculate the pickled mustard greens with LJ036 of *Lactobacillus plantarum*, and add brine with a salt content of 8 wt% until the mustard greens are submerged. The volume of the inoculated live bacteria solution / (live bacteria solution + pickled mustard greens + brine) = 2%. The brine is obtained by adding salt to 1 to 1.5 times the weight of the mustard greens after boiling and cooling water.

[0057] S5. Fermentation: Ferment at 30℃ for 14 days.

[0058] S6. Termination of fermentation: When the total acidity of the mustard greens reaches 8.7g / kg, the greens are taken out to obtain sour mustard greens.

[0059] S7. Preservation of Sour Mustard Greens: Nisin, a preservative, is added to the above-obtained sour mustard greens at a dosage of 0.8 g / kg. After vacuum quantitative packaging, the mixture is sterilized under ultra-high pressure (600 MPa) for 5 minutes and then stored and transported under cold chain at 4℃. The shelf life of this sour mustard green is 6 months.

[0060] Comparative Example 1

[0061] A method for processing pickled mustard greens using natural fermentation includes the following steps:

[0062] S1. Raw material cleaning: After cleaning the mustard greens, treat them with ozone water with a concentration of 5mg / L for 15 minutes to remove pesticide residues, and then air dry to remove surface water.

[0063] S2. Mustard green preparation: Weigh the mustard greens, marinate them with salt at 8% of their weight for 3 hours, pack them into a fermentation container, and press them down layer by layer until the container is 80% full. Then add brine with a salt content of 8wt% (obtained by adding salt to boiled and cooled water) until the mustard greens are submerged.

[0064] S3, Fermentation: Ferment at 30℃ for 24 days.

[0065] S4. Termination of fermentation: When the total acidity of the mustard greens reaches 8.7g / kg, the greens are taken out to obtain sour mustard greens.

[0066] S5. Preservation of Sour Mustard Greens: Add 0.8 g / kg of nisin to the above-obtained sour mustard greens, vacuum-pack them in quantitative quantities, sterilize them under ultra-high pressure of 600 MPa for 5 minutes, and store and transport them in a cold chain at 4℃. The shelf life of this sour mustard greens is 6 months.

[0067] The changes in nitrite content during the fermentation of mustard greens at 30℃ in Example 2 and Comparative Example 1 were investigated. In Example 2, fermentation was not terminated temporarily, and the monitoring time was the same as for natural fermentation. The results are shown in Table 2. The acetoin and total acid content of the sour mustard green products from Example 2 and Comparative Example 1 were also investigated, and the results are shown in Table 3.

[0068] Table 2. Changes in nitrite content during mustard fermentation at 30℃ in Example 2 and Comparative Example 1 (Unit: mg / kg)

[0069]

[0070] Table 3. Acetoin and total acid content of pickled mustard greens products from Example 2 and Comparative Example 1 (unit: mg / kg)

[0071] index Example 2 (inoculation and fermentation for 14 days) Comparative Example 1 (natural fermentation for 24 days) Acetoni (3-hydroxy-2-butanone) 62.0 15.0 Total acidity (calculated as lactic acid) 8.7 8.7

[0072] As shown in Table 2, both Example 2 (fermented mustard greens with Lactobacillus plantarum LJ036) and Comparative Example 1 (naturally fermented mustard greens CK) had peak nitrite levels at 30℃. In Example 2, the peak nitrite level was reached on day 4 after inoculation, with a nitrite content of only 1.26 mg / kg. The nitrite content was very low throughout the fermentation process with Lactobacillus plantarum LJ036, all below 1.3 mg / kg. In contrast, Comparative Example 1 showed a peak nitrite level on day 8 of the pickling process, with a nitrite content as high as 208.8 mg / kg. The nitrite content decreased to below 20 mg / kg on day 22 and was 5.3 mg / kg on day 24.

[0073] As shown in Table 3, the total acid content of Example 2 after 14 days of fermentation was 8.7 mg / kg, while that of Comparative Example 1 after 24 days of fermentation was also 8.7 mg / kg. The inoculation and fermentation time of Example 2 was shortened by 41.7%, and the acetoin content in the fermented product of Example 2 was 4.13 times that of Comparative Example 1. Acetaminophen is an important carbon-based flavor compound in fermented vegetables. Studies have shown that acetoin is related to the transformation of flavor compounds such as diacetyl, acetaldehyde, pyruvic acid, and dimethylglyoxal. The acetoin content, to a certain extent, reflects the metabolism and accumulation of other flavor compounds. For example, diacetyl gives kimchi a creamy aroma, constituting its unique flavor. In summary, the *Lactobacillus plantarum* LJ036 of this invention plays a role in degrading nitrite and enhancing aroma in the fermentation of mustard greens, while also shortening the fermentation time.

[0074] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make many possible variations and modifications to the technical solutions of the present invention using the methods and techniques disclosed above, or modify them into equivalent embodiments with equivalent changes, without departing from the spirit and technical essence of the present invention. Therefore, any simple modifications, equivalent substitutions, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solutions of the present invention shall still fall within the protection scope of the technical solutions of the present invention.

Claims

1. A multifunctional plant lactobacillus strain, characterized in that, The name of the multifunctional plant lactobacillus is Lactiplantibacillus plantarum LJ036, and its accession number at the China Center for Type Culture Collection is CCTCC NO: M 20241710.

2. The application of the multifunctional Lactobacillus plantarum as described in claim 1 in the processing of pickled mustard greens.

3. The application according to claim 2, characterized in that, The application includes the following steps: S1. Raw material cleaning: After the mustard greens are cleaned and pesticide residues are removed, they are dried or shaken to remove surface water. S2. Raw material sterilization treatment: The mustard greens obtained in step S1 are sterilized by pulse ultrasound. S3. Activation of multifunctional plant lactobacillus: The multifunctional plant lactobacillus is activated to obtain a live bacterial solution; S4. Mustard green pickling and inoculation: Weigh the mustard greens, pickle them with 6% to 8% salt by weight of the mustard greens, pack them into a fermentation container and press them down layer by layer, with a filling volume of 70% to 80%. Inoculate the live bacteria solution obtained in step S3 into the pickled mustard greens, and then add salt water with a salt content of 6wt% to 8wt% until the mustard greens are submerged. S5. Fermentation: Control the temperature at 25℃~30℃, and ferment for 12d~16d; S6. Termination of fermentation: When the total acidity of the mustard greens reaches 8.0g / kg~10.0g / kg, the greens are harvested to obtain sour mustard greens; S7. Preservation of pickled mustard greens: The pickled mustard greens obtained in step S6 are preserved using a fence-like anti-corrosion and preservation technology to complete the processing of pickled mustard greens.

4. The application according to claim 3, characterized in that, In step S3, the activation process is as follows: The multifunctional *Lactobacillus plantarum* is first cultured on an MRS solid medium slant, and then cultured on an MRS liquid medium at 36℃~38℃ for 24h~48h. The resulting fermentation broth is centrifuged at 3℃~5℃ and 5000rpm~7000rpm for 10min~15min, and the bacterial cells are collected. The cells are washed 2~3 times with physiological saline, and then resuspended in physiological saline to adjust the bacterial concentration to 10. 7 cfu / mL ~10 8 The cfu / mL concentration was used to obtain a live bacterial culture, which was then refrigerated at 4℃~6℃.

5. The application according to claim 3, characterized in that, In step S4, the pickling time is 2h to 3h, and the volume of the inoculated live bacteria liquid / (live bacteria liquid + pickled mustard greens + brine) = 1% to 2%.

6. The application according to claim 3, characterized in that, In step S7, the preservation process of the fence anti-corrosion and preservation technology is as follows: 0.5g / kg to 1.0g / kg of nisin is added to the pickled mustard greens, vacuum quantitatively packaged, sterilized under 500MPa to 600MPa pressure for 5min to 7min, and cold-chain stored and transported at 4℃ to 6℃; the shelf life is more than 6 months.

7. The application according to any one of claims 3 to 6, characterized in that, In step S1, the process of removing pesticide residues is as follows: the washed mustard greens are treated with high-concentration ozone water, the ozone concentration of the high-concentration ozone water is 5mg / L to 10mg / L, and the treatment time is 10min to 20min.

8. The application according to any one of claims 3 to 6, characterized in that, In step S2, the pulsed ultrasound is either single-frequency pulsed ultrasound or dual-frequency pulsed ultrasound. The frequency of the single-frequency pulsed ultrasound is 80 kHz to 100 kHz, and the frequency of the dual-frequency pulsed ultrasound is 80 kHz to 100 kHz. Each ultrasound treatment lasts for 20 to 30 seconds, with an interval of 5 to 6 seconds, for a total duration of 20 to 30 minutes.

9. The application according to any one of claims 3 to 6, characterized in that, In step S4, the preparation process of the brine is as follows: prepare 1 to 1.5 times the weight of the mustard greens in boiled and cooled water, add salt, and prepare brine.

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