Bacteriostatic lactobacillus plantarum with high bacteriocin yield and application thereof

Abstract

The invention discloses a strain of lactobacillus plantarum, which is named as L.plantarum ZW1, and the preservation number of the lactobacillus plantarum is CGMCC (China General Microbiological Culture Collection Center) 26685. The lactobacillus plantarum provided by the invention is a lactic acid bacterium with relatively strong broad-spectrum inhibition capability, and after the interference of organic acid and hydrogen peroxide is eliminated and protease treatment verification is carried out, the strain is determined to have relatively strong bacteriocin production capability. Probiotic performance evaluation shows that the strain has good digestive tract fluid tolerance, and the L.plantarum CGMCC26685 has good intestinal tract probiotic characteristics, can significantly enrich mouse intestinal flora in an in-vitro intestinal flora fermentation model, increase the relative abundance of the phylum of thick-walled bacteria, reduce the relative abundance of the phylum of proteobacteria, proteusbacillus and shigella, and improve the intestinal tract fluid resistance of mice. The number of flora OTU is obviously higher than that of lactobacillus rhamnosus LGG and lactobacillus plantarum L.plantarum PL4 group which does not produce bacteriocin, and the Chao1 index, the Shannon index and the Simpson index are obviously improved compared with those of a control group. It is shown that the L.plantarum CGMCC26685 has good intestinal tract beneficial activity, and has the potential of further performance excavation and application expansion.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, in particular to an antibacterial plant lactobacillus with high bacteriocin production and application thereof. Background Art

[0002] Lactic acid bacteria (LAB) belong to the family Lactobacillaceae, are facultative anaerobic, Gram-positive, and are one of the generally recognized safe microorganisms. Lactic acid bacteria include multiple genera, such as Lactobacillus and Pediococcus, and most of them are beneficial bacteria with important physiological functions such as regulating physiological functions and improving intestinal ecological balance and function.

[0003] Bacteriocin, also known as antimicrobial peptide, is a protein or biologically active polypeptide produced by bacteria during metabolic activities. These polypeptides or proteins have certain antibacterial activity and can inhibit the growth of pathogenic bacteria. Most bacteria can produce one or more bacteriocins. Bacteriocins are genetically encoded and are not easy to develop drug resistance. They are non-toxic to animals at most effective antibacterial concentrations. Their strong antibacterial activity and natural harmlessness make bacteriocins a potential substitute for antibiotics.

[0004] Bacteriocin-producing lactic acid bacteria can stimulate the growth of beneficial microorganisms and inhibit the growth and reproduction of harmful or pathogenic microorganisms such as Escherichia coli, Staphylococcus aureus, Listeria monocytogenes, and Shigella, regulate the host's defense mechanism, form colonization in the colon, and enhance the availability of nutrients. Bacteriocin-producing lactic acid bacteria can affect the intestinal microbiota through competition for nutrients or adhesion sites, change the composition of the intestinal microbiota, and improve the intestinal microenvironment. Bacteriocin-producing lactic acid bacteria can not only regulate the intestinal flora to correct ecological imbalance, but also serve as natural antibacterial compounds to treat or maintain some important human diseases, such as colon infection, inflammatory bowel disease, obesity, and colorectal cancer. Therefore, further exploring bacteriocin-producing lactic acid bacteria with efficient antibacterial effects and analyzing and evaluating their intestinal beneficial activities will play an important role in promoting the application of lactic acid bacteria in regulating intestinal microecology and intervening or treating diseases. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides an antibacterial Lactobacillus plantarum with high bacteriocin production and application thereof.

[0006] The present invention is achieved through the following technical solutions:

[0007] The first object of the present invention is to provide an antibacterial Lactobacillus plantarum with a high yield of bacteriocin, the strain is Lactobacillus plantarum ZW1, and its classification name is Lactobacillus plantarum (Lactobacillus plantarum). The strain has been deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on February 27, 2023, and the deposit address is the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Beijing, and its deposit number is CGMCC No. 26685.

[0008] Furthermore, the antibacterial Lactobacillus plantarum is prepared by the following method: taking a fermented food sample, diluting and separating it with MRS culture medium, selecting white colonies that resemble lactic acid bacteria, and finally identifying it as Lactobacillus plantarum (Lactobacillus plantarum) through morphological observation, physiological and biochemical characteristics identification, and phylogenetic tree homology analysis, and naming it Lactobacillus plantarum ZW1.

[0009] The plant lactobacillus ZW1 has the following bacterial characteristics:

[0010] (1) Colony morphology: round, milky white, opaque, about 1-2 mm in diameter, with smooth edges and a moist, shiny surface;

[0011] (2) Microscopic morphology: short rod-shaped, Gram-positive, without flagella, and does not produce spores;

[0012] (3) Growth characteristics: The optimal fermentation pH is 5.5 and the temperature is 37°C. The fermentation cycle is short, and it enters the logarithmic growth phase after 4 hours and enters the stable phase after 12 hours.

[0013] The Lactobacillus plantarum ZW1 provided by the present invention has the following probiotic characteristics:

[0014] (1) The cells can survive in artificial gastric fluid at pH 2.0 for 3 h with a survival rate of over 80%;

[0015] (2) After being treated with artificial gastric juice, the cells were able to survive in artificial intestinal juice at pH 6.8 for 4 h with a survival rate of over 60%;

[0016] (3) The cells can survive in MRS medium with a bile salt concentration of 0.03% for 24 h with a survival rate of more than 70%.

[0017] (4) It has a significant inhibitory effect on conditionally pathogenic bacteria Escherichia coli and Staphylococcus epidermidis, with the diameters of the inhibition zones being approximately 25 mm and 23 mm, respectively.

[0018] (5) Compared with the experimental groups supplemented with the model probiotic Lactobacillus rhamnosus LGG and the non-bacteriocin-producing Lactobacillus plantarum PL4, the addition of high-concentration ZW1 significantly enriched the species composition of the mouse fecal flora, the number of OTUs in the flora significantly increased, the biodiversity-related index significantly improved, the relative abundance of beneficial bacteria significantly increased, and the relative abundance of harmful bacteria significantly decreased.

[0019] The second object of the present invention is to provide a bacteriocin produced by the Lactobacillus plantarum.

[0020] The third object of the present invention is to provide the use of the plant lactobacillus or its bacterial suspension or its fermentation liquid in the preparation of antibacterial products.

[0021] Furthermore, the antibacterial bacteria are selected from Escherichia coli, Staphylococcus epidermidis, Streptococcus, Pseudomonas aeruginosa, Salmonella, Shigella, Campylobacter jejuni, and Helicobacter pylori.

[0022] The fourth object of the present invention is to provide the use of the bacteriocin in the preparation of antibacterial products.

[0023] Furthermore, the antibacterial bacteria are selected from Escherichia coli, Staphylococcus epidermidis, Streptococcus, Pseudomonas aeruginosa, Salmonella, Shigella, Campylobacter jejuni, and Helicobacter pylori.

[0024] The fifth object of the present invention is to provide a probiotic agent, comprising the plant lactobacillus or its bacterial suspension or fermentation liquid.

[0025] The sixth object of the present invention is to provide the use of probiotics in the preparation of foods and medicines having the function of regulating intestinal flora.

[0026] The seventh object of the present invention is to provide a method for determining the antibacterial substance produced by the plant lactobacillus, comprising the following steps: by eliminating organic acid interference, eliminating hydrogen peroxide interference, and protein sensitivity inhibition ability testing, determining that the antibacterial substance of the plant lactobacillus is a bacteriocin in the fermentation supernatant.

[0027] Further, the method for eliminating the interference of organic acid is as follows: the pH of the supernatant of the fermentation broth of the tested Lactobacillus plantarum is measured, the pH is adjusted to 4-7 with a NaOH solution, and then an antibacterial experiment is performed, and a blank culture medium adjusted to the same pH with lactic acid and acetic acid is used as a control;

[0028] The method for eliminating the interference of hydrogen peroxide is as follows: using the fermentation supernatant of the untreated Lactobacillus plantarum as a blank control, adding catalase to the sterile supernatant of the Lactobacillus plantarum to evaluate the change in the antibacterial activity of the fermentation supernatant after treatment;

[0029] The method for protease sensitivity comprises adjusting the fermentation supernatant of the tested plant lactobacillus to pH 1.9-2.1 with acid, adding 1 mg / mL of pepsin, reacting for a certain time, then killing the enzyme, adjusting the pH to 4-7, and adding catalase to measure the antibacterial activity.

[0030] The method for determining the antibacterial substance produced by plant lactobacillus specifically comprises the following steps:

[0031] (1) Eliminate organic acid interference: Measure the pH of the supernatant of the test lactic acid bacteria fermentation broth, adjust the pH to 5.0 with 2 mol / L NaOH solution, and then perform the antibacterial experiment. Use a blank culture medium adjusted to the same pH with 1 mol / L lactic acid or acetic acid as a control.

[0032] (2) Eliminate interference from hydrogen peroxide: Using the untreated fermentation supernatant of the test bacteria as a blank control, add 1 mg / mL of catalase to the sterile supernatant of the test lactic acid bacteria and evaluate the changes in the antibacterial activity of the fermentation supernatant after treatment.

[0033] (3) Protease sensitivity: The pH of the fermentation supernatant of the test lactic acid bacteria was adjusted to 2.0 with 1 mol / L HCl, and 1 mg / mL of pepsin was added. The supernatant was placed in a 37°C metal bath for 2 h and then boiled to inactivate the enzyme. The pH was then adjusted to 5.0, and catalase was added at a final concentration of 1 mg / mL for the antibacterial activity determination.

[0034] The above technical solution of the present invention has the following advantages compared with the prior art:

[0035] The Lactobacillus plantarum ZW1 provided by the present invention has a strong inhibitory effect on conditional pathogenic bacteria Escherichia coli and Staphylococcus epidermidis, has a good intestinal probiotic effect, and has a high ability to resist gastrointestinal stress and bile salt. In addition, the strain has the characteristics of fast growth rate and short fermentation cycle, and has the potential for further performance mining and application expansion.

[0036] The Lactobacillus plantarum ZW1 described in the present invention can significantly enrich the intestinal flora, significantly increase the relative abundance of Firmicutes, and reduce the relative abundance of Proteobacteria, Proteus and Shigella. The OTU number of the flora is significantly higher than that of the experimental group with the addition of the probiotic Lactobacillus rhamnosus LGG and the non-bacteriocin-producing Lactobacillus plantarum PL4, and the Chao1, Shannon and Simpson indexes related to biodiversity are significantly improved compared with the control group. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] In order to make the content of the present invention more clearly understood, the present invention is further described in detail below according to specific embodiments of the present invention in conjunction with the accompanying drawings, wherein:

[0038] Figure 1 It is a colony morphology diagram of Lactobacillus plantarum ZW1 of the present invention.

[0039] Figure 2 It is the growth curve and pH change curve of Lactobacillus plantarum ZW1 of the present invention.

[0040] Figure 3 The present invention shows the antibacterial activity of Lactobacillus plantarum ZW1 against Escherichia coli.

[0041] Figure 4 The present invention shows the antibacterial activity of Lactobacillus plantarum ZW1 against Staphylococcus epidermidis.

[0042] Figure 5 It is the tolerance of Lactobacillus plantarum ZW1 of the present invention to artificial gastrointestinal fluid.

[0043] Figure 6 It is the bile salt tolerance of Lactobacillus plantarum ZW1 of the present invention.

[0044] Figure 7 It is the alpha diversity index analysis of the present invention, wherein a: Chao1 index; b: observed OUT number; c: Shannon index; d: Simpson index.

[0045] Figure 8 This is the OTU distribution of intestinal microorganisms of the present invention: a: adding different concentrations of L. plantarum ZW1 for comparative analysis; b: adding different concentrations of L. rhamnosus GG for comparative analysis; c: adding different concentrations of L. plantarum PL4 for comparative analysis; d: adding low concentration of lactic acid bacteria for comparison; e: adding high concentration of lactic acid bacteria.

[0046] Fig. 9 This is the relative abundance of species at the genus level of the present invention. DETAILED DESCRIPTION

[0047] The present invention is further described below in conjunction with the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it, but the embodiments are not intended to limit the present invention.

[0048] The experimental methods in the following examples are conventional methods unless otherwise specified. The raw materials involved in the following examples are common commercial products and can be purchased from the market unless otherwise specified.

[0049] The present invention is further described in detail below in conjunction with the drawings and embodiments:

[0050] Example 1: Isolation and identification of Lactobacillus plantarum ZW1 of the present invention

[0051] 1) Strain Isolation

[0052] Take 1g of fermented pear juice sample and add it to 9mL of normal saline, shake it to mix it thoroughly, and take 10 -5 , 10 -6 , 10 -7 The bacterial solution was evenly spread on MRS plates and cultured in an anaerobic incubator at 37°C. After 24 hours, the strains were preliminarily identified by colony morphology and microscopic examination, and strains that met the characteristics were selected for streaking purification and culture.

[0053] MRS medium formula (g / L): glucose 20, beef extract 10, peptone 10, yeast extract 5, anhydrous sodium acetate 5, diammonium citrate 2, potassium dihydrogen phosphate 2, manganese sulfate 0.19, magnesium sulfate 0.58, Tween 80 1mL, agar powder 15, pH 6.5.

[0054] 2) Morphology and 16S rDNA identification of strains

[0055] The colony morphology of the plant lactobacillus ZW1 of the present invention is shown in Figure 1 The colonies are round, milky white, opaque, about 1 to 2 mm in diameter, with smooth edges and a moist, shiny surface.

[0056] DNA extraction and 16S rDNA identification were performed on the selected target strains. The 16S rDNA fragment was amplified by PCR using universal primers 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' and 1492R: 5'-GGTTACCTTGTTACGACTT-3'. 1.0% gel agar electrophoresis was used to detect the DNA extraction and the size of the PCR amplification product. The obtained target fragments were sequenced and analyzed, and the sequencing results were compared in the NCBI database. Combined with homology analysis, the obtained strain was determined to be Lactobacillus plantarum, and its classification was named Lactobacillus plantarum. It was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on February 27, 2023. The deposit address is the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No. 26685.

[0057] Example 2: Growth curve and pH curve of Lactobacillus plantarum ZW1 of the present invention

[0058] The activated seed liquid of Lactobacillus plantarum ZW1 was inoculated with 1% of the inoculum into 100 mL of MRS liquid medium under sterile conditions and cultured anaerobically at 37°C for 24 h. Samples were taken every 4 h to measure the OD 600 Each sample was tested three times, with the incubation time as the horizontal axis and the OD as the horizontal axis. 600Or pH value as the vertical axis, draw the growth curve and pH curve of the bacteria (see Figure 2 ). Lactobacillus plantarum ZW1 entered the logarithmic growth phase at 4h and entered the stable phase after 12h, with a short fermentation cycle. The pH curve showed a slow fluctuation decrease from 0 to 4h and a rapid decrease from 4 to 12h, indicating that this stage was the stage of large-scale acid production by ZW1. After 12h, the pH value remained at around 3.6.

[0059] Example 3: Evaluation of the antibacterial activity of Lactobacillus plantarum ZW1 of the present invention

[0060] The activated seed liquid of Lactobacillus plantarum ZW1 was inoculated at 1% into 20 mL of MRS liquid culture medium under sterile conditions, cultured anaerobically at 37°C for 24 h, the bacterial liquid was collected, centrifuged at 4500 r / min for 15 min, and the supernatant was collected and filtered with a 0.22 μm filter membrane.

[0061] The antibacterial test used the Oxford cup method. First, two indicator bacteria, Escherichia coli and Staphylococcus epidermidis, were activated. About 20 mL of LB agar medium was added to a disposable sterile culture dish and allowed to stand horizontally. After solidification, 0.1 mL of indicator bacteria solution (10 6 CFU / mL), spread evenly; use sterile tweezers to place an Oxford cup (8mm in diameter) on the plate, take 200μL of supernatant and add it to the Oxford cup; place the culture dish forward at 37℃ and culture for 24h, measure the size of the inhibition zone, and repeat three times for each indicator bacteria. Figure 3 and Figure 4 As shown in the figure, when Escherichia coli is used as the indicator bacteria, the inhibition zone is about 25 mm; when Staphylococcus epidermidis is used as the indicator bacteria, the inhibition zone is about 23 mm. The results show that Lactobacillus plantarum ZW1 has a good antibacterial effect.

[0062] Example 4: Confirmation of bacteriocin production by Lactobacillus plantarum ZW1 of the present invention

[0063] (1) Eliminate organic acid interference

[0064] Measure the pH of the supernatant of plant lactobacillus ZW1 fermentation liquid, adjust the pH to 5.0 with 2mol / LNaOH solution, and then carry out antibacterial experiment. The blank culture medium adjusted to the same pH with 1mol / L lactic acid and acetic acid was used as a control. The result shows that the transparent circle produced by the fermentation supernatant with pH of 5.0 on the Escherichia coli growth plate is about 20mm, which is significantly smaller than the original fermentation supernatant, but can still produce a transparent circle of a certain size. In addition, the MRS culture medium with lactic acid and acetic acid added almost does not produce a transparent circle on the indicator bacteria growth plate. The above explanation shows that the substance that has antibacterial effect has other substances in addition to acidic products.

[0065] (2) Eliminate interference from hydrogen peroxide

[0066] The fermentation supernatant of Lactobacillus plantarum ZW1 was used as a blank control, and 1 mg / mL of catalase was added to the sterile supernatant to evaluate the changes in the antibacterial activity of the fermentation supernatant after treatment. The results showed that after adding catalase, the inhibition zone on the Escherichia coli growth plate was about 26 mm, which was no significant difference from the original fermentation supernatant, so the interference of hydrogen peroxide could be ruled out.

[0067] (3) Protease sensitivity

[0068] The fermentation supernatant of Lactobacillus plantarum ZW1 was adjusted to pH 2.0 with 1 mol / L HCl, pepsin 1 mg / mL was added, and the enzyme was boiled and inactivated after being placed in a 37°C metal bath for 2 hours. The pH was then adjusted to 5.0, and catalase was added at a final concentration of 1 mg / mL for antibacterial activity determination. The results showed that after treatment with trypsin and pepsin, basically no inhibition zone was produced on the Escherichia coli growth plate, indicating that the antibacterial substance produced was a protein substance, and it can be preliminarily determined that the antibacterial substance produced by Lactobacillus plantarum ZW1 is bacteriocin.

[0069] Example 5: Tolerance of Lactobacillus plantarum ZW1 of the present invention to artificial gastrointestinal fluid

[0070] Artificial gastric juice formula: Take 0.1 mol / L HCl and adjust the pH to 2.0 with distilled water, add 0.01 g / mL pepsin, and sterilize the microporous filter membrane for use.

[0071] Artificial intestinal fluid formula: dissolve 2.04g of KH2PO4 with appropriate amount of water and make up to 150ml, adjust the pH of the solution to 6.8 with 4% sodium hydroxide, add 0.01g / ml of trypsin, mix well, filter and sterilize for later use.

[0072] The 24h fermentation liquid of Lactobacillus plantarum ZW1 was inoculated into artificial gastric juice at a volume ratio of 1:10, and cultured at a constant temperature of 37°C. Samples were taken at 0, 1, 2, and 3h, and the live bacteria in the artificial gastric juice were diluted and spread for counting; the suspension inoculated in artificial gastric juice for 3h was further added to artificial intestinal juice at a volume ratio of 1:10, and samples were taken and counted at 4, 5, 6, 7, and 8h to obtain the number of live bacteria, and the survival rate was calculated:

[0073] Survival rate / % = (N i / N0)×100%(1), formula (1) evaluates the strain's tolerance to simulated gastrointestinal fluid. i is the number of viable bacteria corresponding to different times; N0 is the number of viable bacteria corresponding to 0h. Figure 5 In artificial gastric juice, the survival rate at 3 h was 83.92%; in artificial intestinal juice, the survival rate at 4 h was 60.07%, indicating that the bacteria has excellent ability to tolerate gastrointestinal juice.

[0074] Example 6: Tolerance of Lactobacillus plantarum ZW1 of the present invention to bile salts

[0075] 0.03%, 0.06%, 0.09% and 0.12% bile salts were added to the MRS medium, respectively, and sterilized for later use. Lactobacillus plantarum ZW1 was inoculated into bile salt medium of different concentrations, with MRS medium as control, and cultured at 37°C for 24 hours. Samples were taken every 8 hours for viable bacteria count, and the survival rate was calculated according to formula (1). The results are shown in Figure 6 As shown, when the bile salt concentration was 0.03%, the survival rate of the strain reached 72.1% after 24 hours, and when the bile salt concentration was 0.12%, the survival rate of the strain was still close to 50% after 24 hours, indicating that the bacteria have a certain ability to tolerate bile salts.

[0076] Example 7: Evaluation of intestinal probiotic performance of Lactobacillus plantarum ZW1 of the present invention

[0077] Normal mouse feces were homogenized in 0.1 mol / L PBS solution at a volume ratio of 10%, and then inoculated into the basal medium at a final concentration of 10% as the fermentation medium. The target strain, the model probiotic Lactobacillus rhamnosus LGG, and a non-bacteriocin-producing L. plantarum PL4 were activated, and 1 mL of 5×10 9 and 5×10 8 CFU / mL of fermentation broth was added to the fermentation medium and cultured anaerobically at 37°C for 48 h. The experimental groups were shown in the table. The sample precipitate after fermentation was analyzed after 16srDNA amplicon sequencing.

[0078] Table 1 Experimental groups

[0079]

[0080]

[0081] Simpson, Shannon, Observed species and Chao1 are the main components of the α diversity index. The Simpson and Shannon indices are proportional to the diversity of the bacterial community, while Observed species and Chao1 represent the number of operational taxonomic units (OTUs) at this sequencing depth, which can reflect the species richness in the bacterial community. Figure 7The results showed that compared with the control group, the number of observed species, Chao1 diversity index, Shannon index and Simpson index of the low-dose LGG group and the low-dose LPPL4 group did not change significantly, indicating that the addition of low-concentration LGG and L. plantarum PL4 had no significant effect on the richness and diversity of the fecal flora of mice. However, the number of observed species, Chao1 diversity index, Shannon index and Simpson index of the high-dose LGG group, the high-dose LPPL4 group and the low-dose LPZW1 group were improved compared with the negative control group, indicating that the high-concentration LGG, L. plantarum PL4 and the low-dose LPZW1 had the same effect on the fecal flora of mice, and both increased the richness and diversity of the flora. The number of observed species, Chao1 diversity index, Shannon index and Simpson index of the high-dose LPZW1 group were significantly improved compared with the NC group, indicating that the addition of high-concentration bacteriocin-producing L. plantarum ZW1 significantly enriched the flora in the feces of mice.

[0082] OTU statistics were performed for all groups. First, the fecal flora of the NC group was compared with those of the groups supplemented with different concentrations of LGG, L. plantarum PL4, and L. plantarum ZW1 ( Figure 8 a, 8b, 8c), and found that the OTUs specific to each lactic acid bacteria increased with the increase of the added concentration. The fecal flora of different lactic acid bacteria added at the same concentration were compared and analyzed ( Figure 8 d, 8e), and found that when adding low concentrations, among the three groups, the fecal flora with L. plantarum ZW1 had the most unique OTUs, reaching 231, which was higher than the 113 with LGG and the 104 with L. plantarum PL4; when adding high concentrations, the unique OUTs of the fecal flora with L. plantarum ZW1 were very close to those with LGG, and significantly higher than those with L. plantarum PL4, indicating that the bacteriocin-producing Lactobacillus L. plantarum ZW1, like the recognized intestinal probiotics, can significantly increase the richness of fecal flora.

[0083] At the genus level, Fig. 9It can be seen that compared with the control group, the relative abundance of Bacteroides, Parabacteroides, Clostridium_sensu_stricto_13 and Lactobacillus in other groups increased, and the high-dose LPZW1 group had Methylobacter that was not in other groups, while the relative abundance of intestinal harmful bacteria Escherichia-Shigella and Proteus decreased, among which the LPZW1H group had the largest decrease, by 11.3% and 1.5%, respectively. The above shows that ZW1 can improve the composition of intestinal microorganisms.

[0084] Obviously, the above embodiments are merely examples for clear explanation and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the protection scope of the invention.

Claims

1. A strain of Lactobacillus plantarum, characterized in that The plant lactobacillus is named L. plantarum ZW1. The plant lactobacillus was deposited on February 27, 2023 at the General Microbiology Center of the China Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Beijing, and its deposit number is CGMCC No. 26685.

2. A bacteriocin, characterized in that Produced by the plant lactobacillus described in claim 1.

3. Application of plant lactobacillus or its bacterial suspension or its fermented liquid according to claim 1 in preparing an antibacterial product.

4. According to the use described in claim 3, the antibacterial bacteria are selected from Escherichia coli, Staphylococcus epidermidis, Streptococcus, Pseudomonas aeruginosa, Salmonella, Shigella, Campylobacter jejuni, and Helicobacter pylori.

5. Use of the bacteriocin described in claim 2 in the preparation of antibacterial products.

6. The use according to claim 5, wherein the antibacterial bacteria are selected from Escherichia coli, Staphylococcus epidermidis, Streptococcus, Pseudomonas aeruginosa, Salmonella, Shigella, Campylobacter jejuni, and Helicobacter pylori.

7. A probiotic, characterized in that: The invention comprises the plant lactobacillus or its bacterial suspension or fermentation liquid as described in claim 1.

8. Use of the probiotic agent according to claim 7 in the preparation of food and medicine having the function of regulating intestinal flora.

9. A method for determining the antibacterial substance produced by the Lactobacillus plantarum according to claim 1, characterized in that: The following steps are involved: By eliminating the interference of organic acids and hydrogen peroxide, as well as protein sensitivity inhibition ability test, it is determined that the antibacterial substance of Lactobacillus plantarum described in claim 1 is the bacteriocin in the fermentation supernatant.

10. The determination method according to claim 9, characterized in that: in, The method for eliminating organic acid interference is as follows: the pH of the supernatant of the fermentation broth of the tested Lactobacillus plantarum is measured, the pH is adjusted to 4-7 with a NaOH solution, and then an antibacterial experiment is performed, and a blank culture medium adjusted to the same pH with lactic acid and acetic acid is used as a control; The method for eliminating the interference of hydrogen peroxide is as follows: using the fermentation supernatant of the untreated Lactobacillus plantarum as a blank control, adding catalase to the sterile supernatant of the Lactobacillus plantarum to evaluate the change in the antibacterial activity of the fermentation supernatant after treatment; The method for protease sensitivity comprises adjusting the fermentation supernatant of the tested plant lactobacillus to a pH of 1.9-2.1 with acid, adding 1 mg / mL of pepsin, reacting for a certain time, then killing the enzyme, adjusting the pH to 4-7, and adding catalase to measure the antibacterial activity.

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