Antibacterial composition comprising plantaricin peptide derived from lactiplantibacillus plantarum km2 strain as active ingredient

Abstract

The present invention relates to an antibacterial composition comprising, as an active ingredient, a plantaricin peptide derived from a Lactiplantibacillus plantarum KM2 strain, wherein the plantaricin peptide derived from the strain and a plantaricin peptide combination were found to have antimicrobial activity against at least one selected from the group consisting of Flavobacterium sp, Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, and acromobacter xylosoxidans (Alcaligenese xylosoxidans), Salmonella enterica, and Vibrio parahaemolyticus, and the plantaricin peptide and the plantaricin peptide combination were found to have antibacterial activity and a cell wall lysis effect against the Flavobacterium sp. strain, and accordingly, the plantaricin peptide derived from the Lactiplantibacillus plantarum KM2 strain and the plantaricin peptide combination are provided as antimicrobial agents.

Description

Antibacterial composition comprising plantaricin peptide derived from Lactibacillus plantarum KM2 strain as an active ingredient

[0001] The present invention relates to an antibacterial composition comprising, as an active ingredient, a plantaricin peptide derived from Lactiplantibacillus plantarum KM2 strain.

[0002] Lactiplantibacillus plantarum is a microorganism commonly found in fermented foods, such as fermented milk, sausages, cheese, and vegetables. Lb. plantarum is used as a starter culture for fermented foods, such as yogurt and sourdough. It is also known as a probiotic strain that influences immune system regulation and is used to treat gastrointestinal diseases. Most importantly, Lb. plantarum is a food-grade bacterium that has been used in fermented foods for a long time. Some Lb. plantarum strains produce bacteriocins, some of which exhibit antifungal activity. Bacteriocins are peptides, proteins, or complexes that inhibit the growth of bacteria, including both Gram-negative and Gram-positive bacteria. Most bacteriocins exhibit a narrow spectrum of inhibitory activity, but some have been shown to inhibit a broad spectrum of bacteria. The bacteriocin gene operon contains genes for immune proteins, so they do not harm the bacteria themselves. Bacteriocins can inhibit food spoilage and pathogenic bacteria in the food industry. Their use as natural food preservatives provides consumers with a perceived sense of safety.

[0003] Lb. plantarumKM2 was isolated from aged Korean beef, and showed antibacterial activity against seven foodborne pathogens and spoilage bacteria. Genomic analysis confirmed the presence of the bacteriocin operon, plantaricin, in this strain. In this experiment, we aimed to verify the antibacterial activity of a synthetic antibacterial substance based on the bacteriocin gene of Lb. plantarumKM2, determine its antibacterial spectrum, and elucidate the antibacterial mechanism of the synthesized antibacterial substance.

[0004] [Prior Art Literature]

[0005] [Patent Document]

[0006] Korean Patent Publication No. 10-2023-0000719 (Published on January 3, 2023)

[0007] The purpose of the present invention is to provide an antibacterial composition comprising, as an active ingredient, a plantaricin peptide derived from the Lactiplantibacillus plantarum KM2 strain deposited under the accession number KCTC 14637BP.

[0008] The present invention provides an antibacterial composition comprising, as an active ingredient, a plantaricin peptide derived from Lactiplantibacillus plantarum KM2 strain.

[0009] In addition, the present invention provides a health functional food composition comprising the antibacterial composition as an active ingredient.

[0010] In addition, the present invention provides a cosmetic composition comprising the antibacterial composition as an active ingredient.

[0011] In addition, the present invention provides a food preservative composition comprising the antibacterial composition as an active ingredient.

[0012] In addition, the present invention provides an oral therapeutic microbiome pharmaceutical composition comprising the above antibacterial composition as an active ingredient.

[0013] In addition, the present invention provides an oral microbiome regulating composition comprising the antibacterial composition as an active ingredient.

[0014] According to the present invention, it was confirmed that a combination of plantaricin derived from Lactiplantibacillus plantarum KM2 strain deposited under accession number KCTC 14637BP and a synthetic plantaricin peptide exhibits antibacterial activity against at least one strain selected from the group consisting of Flavobacterium sp., Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, Alcaligenes xylosoxidans, Salmonella enterica, and Vibrio parahaemolyticus, and the plantaricin peptide and the combination of the plantaricin peptide As it has been confirmed that it has antibacterial activity and a cell wall dissolving effect against Flavobacterium sp. strains, plantaricin peptides and combinations of plantaricin peptides derived from Lactiplantibacillus plantarum KM2 strains can be provided as antibacterial agents.

[0015] Figure 1 shows the results of comparative analysis of the genes of the plantaricin operon of Lactiplantibacillus plantarum strains.

[0016] Figure 2 shows the results of analyzing the antibacterial activity of synthetic plantaricin against Flavobacterium sp. using Raman spectroscopy.

[0017] Figure 3 shows the results of analyzing the antibacterial activity of synthetic plantaricin against Flavobacterium sp. using a transmission electron microscope.

[0018] The terms used in this specification have been selected from widely used, current terms, taking into account the functions of the present invention. However, these terms may vary depending on the intentions of those skilled in the art, precedents, the emergence of new technologies, etc. Furthermore, in certain cases, terms may be arbitrarily selected by the applicant, and in such cases, their meanings will be described in detail in the relevant description of the invention. Therefore, the terms used in this invention should not be defined simply as names, but rather based on their inherent meanings and the overall content of the present invention.

[0019] Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with their meaning in the context of the relevant technology, and shall not be interpreted in an idealized or overly formal sense unless explicitly defined herein.

[0020] Hereinafter, the present invention will be described in more detail.

[0021] The present invention provides an antibacterial composition comprising, as an active ingredient, one or more plantaricin peptides derived from Lactiplantibacillus plantarum KM2 strain.

[0022] The above Lactiplantibacillus plantarum KM2 strain is a strain deposited under the accession number KCTC 14637BP.

[0023] The above plantaricin peptide may comprise any one amino acid sequence selected from SEQ ID NOs: 1 to 5. The above antibacterial composition may comprise one or more peptides consisting of any one amino acid sequence selected from SEQ ID NOs: 1 to 5.

[0024] Preferably, the antimicrobial composition according to the present invention may be a combination of a plantaricin peptide comprising an amino acid sequence consisting of SEQ ID NO: 1 and a plantaricin peptide comprising at least one amino acid sequence selected from SEQ ID NOs: 2 to 5.

[0025] The above antibacterial composition may exhibit antibacterial activity against at least one strain selected from the group consisting of Flavobacterium sp., Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, Alcaligenes xylosoxidans, Salmonella enterica, and Vibrio parahaemolyticus.

[0026] In addition, the present invention provides a health functional food composition comprising the antibacterial composition as an active ingredient.

[0027] The above food composition can be generally used as a commonly used food.

[0028] The food composition of the present invention can be used as a health functional food. The term "health functional food" refers to a food manufactured and processed using raw materials or ingredients with functional properties beneficial to the human body, as defined by the Health Functional Food Act. "Functionality" refers to ingestion for the purpose of obtaining beneficial effects for health purposes, such as regulating nutrients for the structure and functions of the human body or physiological effects.

[0029] The food composition of the present invention may include conventional food additives, and its suitability as the "food additive" is determined by the specifications and standards for the relevant item in accordance with the general provisions and general test methods of the Food Additive Code approved by the Ministry of Food and Drug Safety, unless otherwise specified.

[0030] Items listed in the above "Food Additives Code" include, for example, chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamic acid; natural additives such as persimmon pigment, licorice extract, crystalline cellulose, high-molecular-weight pigment, and guar gum; and mixed preparations such as sodium L-glutamate preparations, alkaline agents for noodles, preservative preparations, and tar color preparations.

[0031] The food composition of the present invention can be manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc.

[0032] For example, a health functional food in capsule form may include at least one of a hard capsule and a soft capsule. The hard capsule may be prepared by mixing the composition according to the present invention with additives such as excipients and filling a conventional hard capsule, and the soft capsule may be prepared by mixing the composition according to the present invention with additives such as excipients and filling a capsule base such as gelatin. The soft capsule may additionally contain a plasticizer such as glycerin or sorbitol, a coloring agent, a preservative, etc., as needed.

[0033] The definitions of terms for the above excipients, binders, disintegrants, lubricants, flavoring agents, etc. are described in literature known in the art and include those with identical or similar functions. There are no specific restrictions on the type of food, and all health functional foods in the conventional sense are included.

[0034] In addition, the present invention provides a cosmetic composition comprising the antibacterial composition as an active ingredient.

[0035] The above cosmetic composition may contain the above antibacterial composition in an amount capable of exhibiting antibacterial activity.

[0036] The above cosmetic composition can be manufactured in the form of a general emulsified formulation or a solubilized formulation using a commonly known manufacturing method. The above cosmetic composition can be appropriately selected depending on the purpose, and can be formulated into one or more formulations selected from the group consisting of an emollient toner, an astringent toner, a nourishing toner, an eye cream, a nourishing cream, a cleansing cream, a cleansing foam, a cleansing water, a powder, an essence, a pack, etc.

[0037] In addition, the present invention provides a food preservative composition comprising the antibacterial composition as an active ingredient.

[0038] The above preservatives refer to agents added for the purpose of preserving food, and therefore must not be harmful to the human body and must not impair the quality of the food due to their addition.

[0039] In addition, the present invention provides an oral therapeutic microbiome pharmaceutical composition comprising the above antibacterial composition as an active ingredient.

[0040] The above microbiome pharmaceutical composition exhibits a preventive or therapeutic effect against infectious enteric diseases caused by harmful bacteria. The harmful bacteria may be at least one strain selected from the group consisting of Flavobacterium sp., Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, Alcaligenes xylosoxidans, Salmonella enterica, and Vibrio parahaemolyticus.

[0041] The pharmaceutical composition may be prepared by further including a pharmaceutically acceptable adjuvant. The adjuvant may be at least one selected from excipients, disintegrants, sweeteners, binders, coating agents, bulking agents, lubricants, glidants, or flavoring agents. The pharmaceutical composition may be formulated by including at least one pharmaceutically acceptable carrier and diluent. The pharmaceutical composition may be in the form of granules, powders, tablets, coated tablets, capsules, suppositories, liquids, syrups, juices, suspensions, emulsions, drops, or injectable solutions. The pharmaceutically acceptable carriers and diluents include, but are not limited to, excipients such as starch, sugar and mannitol, fillers and bulking agents such as calcium phosphate, cellulose derivatives such as carboxymethylcellulose and hydroxypropylcellulose, binders such as gelatin, alginates and polyvinyl pyrrolidone, lubricants such as talc, calcium stearate, hydrogenated castor oil and polyethylene glycol, disintegrants such as povidone and crospovidone, and surfactants such as polysorbates, cetyl alcohol and glycerol.

[0042] The pharmaceutical composition of the present invention can be administered orally according to the intended method. In the case of oral administration, it can be formulated as tablets, troches, lozenges, aqueous suspensions, oily suspensions, prepared powders, granules, emulsions, hard capsules, soft capsules, syrups, elixirs, etc.

[0043] The dosage of the pharmaceutical composition of the present invention may vary depending on the patient's condition, weight, age, sex, health condition, dietary constitution, nature of the preparation, degree of disease, administration time of the composition, administration method, administration period or interval, excretion rate, and drug form, and may be appropriately selected by a person skilled in the art. For example, the dosage may be in the range of about 0.1 to 10,000 mg / kg, but is not limited thereto, and may be administered once or several times a day in divided doses.

[0044] The pharmaceutically effective amount and effective dosage of the pharmaceutical composition of the present invention may vary depending on the formulation method, administration method, administration time, administration route, etc. of the pharmaceutical composition. Those skilled in the art can easily determine and prescribe an effective dosage for the desired treatment. The pharmaceutical composition of the present invention may be administered once a day or divided into several doses.

[0045] In addition, the present invention provides an oral microbiome regulating composition comprising the antibacterial composition as an active ingredient.

[0046] The above microbiome-regulating composition exhibits the effect of improving intestinal health by improving the intestinal microbiome environment by killing harmful bacteria in the intestines. The harmful bacteria may be any one or more strains selected from the group consisting of Flavobacterium sp., Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, Alcaligenes xylosoxidans, Salmonella enterica, and Vibrio parahaemolyticus.

[0047] Hereinafter, to aid understanding of the present invention, experimental examples and examples will be described in detail. However, the following experimental examples and examples are merely illustrative of the present invention and are not intended to limit the scope of the present invention. The experimental examples and examples of the present invention are provided to more fully explain the present invention to those with average knowledge in the art.

[0048] <Experimental Example> Experimental Materials and Methods

[0049] The following experimental examples are intended to provide experimental examples commonly applied to each embodiment according to the present invention.

[0050] 1. Strains and culture conditions

[0051] The Lactiplantibacillus plantarum KM2 strain used in the present invention was isolated from unaged Korean beef and Korean beef aged at 4℃ for 60 days at a restaurant in Gongju-si, Chungcheongnam-do. The Lb. plantarum KM2 strain was deposited at the Korea Center for Biological Resources (KCTC) of the Korea Research Institute of Bioscience and Biotechnology on July 14, 2021, and was assigned the accession number KCTC 14637BP. The Lb. plantarum KM2 strain was used in the experiment after being cultured for 24 hours in MRS medium (Becton, Dickinson and Co., USA) at 30℃.

[0052] 2. Genome Comparative Analysis

[0053] The complete genome sequence of Lb. plantarumKM2 strain is registered in GenBank (Accession numbers: CP069282). For comparative genomic analysis of the plantarisin operon of Lb. plantarumKM2 strain, Lb. plantarum8P-A3(GCA_004403045.1), Lb. plantarumB21(GCA_000931425.2), Lb. plantarumFBL-3a(GCA_003999275.1), Lb. plantarumKACC 92189(GCA_003692595.1), Lb. plantarumLL441(GCA_001754025.1), Lb. plantarumMF1298(GCA_001880185.2), Lb. plantarumKACC 92189(GCA_003692595.1), Lb. plantarumLL441(GCA_001754025.1), Lb. plantarumMF1298(GCA_001880185.2), Lb. plantarumKM2 strain were used. Genome sequence data for L. plantarum NCIMB 700965 (GCA_003611015.1) and L. plantarum Q7 (GCA_003999605.1) strains were obtained from the National Center for Biotechnology Information (NCBI) database (http: / / ncbi.nlm.nih.gov / genomes). The sequence homology of conserved genes was assessed using the Basic Local Alignment Search Tool (BLAST), and comparative genome analysis was performed using EDGAR 3.2 (EDGAR; https: / / edgar3.computational.bio.uni-giessen.de) and RAST (Rapid Annotation using Subsystem Technology) (https: / / rast.nmpdr.org / ).

[0054] 3. Prediction of the signal sequence and structure of plantaricin

[0055] The amino acid sequences of all plantaricins, including cleavage sites and signal sequences, were analyzed using SignalP 5.0 (Almagro Armenteros et al., 2019). The tertiary structure of the synthetic plantaricins was predicted using AlphaFold (Jumper et al., 2021).

[0056] 4. Peptide synthesis

[0057] Plantarisin peptides lacking signal peptides (PlnA, E, F, J, and K) were synthesized by Anygen (Gwangju, Korea). The synthesized peptides were purified by high-performance liquid chromatography (HPLC) using a C-18 reversed-phase column, and the masses of all peptides were confirmed by matrix-assisted laser desorption / ionization-time of flight (MALDI-TOF) mass spectrometry (Shimadzu).

[0058] 5. Measurement of antibacterial activity

[0059] The antibacterial activity of the synthesized plantaricin was measured using the disk diffusion method against nine foodborne pathogens (Bacillus cereus KCCM (Korean Culture Center of Microorganisms) 11341, Enterococcus faecalis KCTC 2011, Listeria monocytogenes ATCC (American Type Culture Collection) 19111, Staphylococcus aureus ATCC 12692, Alcaligenes xylosoxidans KCCM 40240, Escherichia coli O157:H7 EDL 933, Flavobacterium sp. KCCM 11374, Salmonella enterica KCCM 11862, and Vibrio parahaemolyticus KCTC 2729). The indicator strains were cultured in Tryptic Soy Broth (TSB) (Becton, Dickinson and Co.) medium. Each indicator strain was inoculated into TSB medium at 1% (v / v) and the wavelength was 600 nm (OD 600 ) in OD 600 = 1.0, the cultured strains were plated on tryptic soy agar (TSA) (Becton, Dickinson and Co.). Synthetic plantaricin was prepared by diluting to 1 nmol / L. Sterilized disks were placed on solid medium, and 20 μL of each peptide was dropped onto the disk. Antibacterial activity was determined by the formation of a clear ring around the disk.

[0060] 6. Minimum inhibitory concentration (MIC) evaluation

[0061] To determine the minimum inhibitory concentration (MIC) of synthetic plantaricin against Flavobacterium sp. KCCM 11374, Flavobacterium sp. KCCM 11374 was inoculated into TSB, and OD 600 When the concentration reached 0.5, synthetic plantaricins, including spPlnA (Spontaneous plantaricin A), spPlnAEJ (Spontaneous plantaricin AEJ), spPlnAEK (Spontaneous plantaricin AEK), spPlnAFJ (Spontaneous plantaricin AFJ), and spPlnAFK (Spontaneous plantaricin AFK), were added at a final concentration range of 0.1 to 1.5 nmol / L. The growth of Flavobacterium sp. KCCM 11374 was measured at 37°C for 24 h using a Synergy HT plate reader (BioTek, Winooski, VT, USA). All measurements were repeated three times with independent cultures.

[0062] 7. Raman spectrum

[0063] After treatment of Flavobacterium sp. KCCM 11374 with synthetic plantaricin, changes in cellular metabolism were analyzed using Raman spectroscopy (RAMANtouch, Nanophoton Co., Osaka, Japan). Principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) were performed based on the data.

[0064] 8. Transmission electron microscope (TEM) analysis

[0065] Changes in the cell wall of Flavobacterium sp. KCCM 11374 treated with synthetic plantaricin were observed using TEM (JEM-1010 TEM).

[0066] Example 1. Plantaricin operon of Lb. plantarumKM2

[0067] Genome analysis of the Lb. plantarum KM2 strain confirmed the plantaricin operon, known to produce bacteriocins. Comparison with the genomes of other Lb. plantarum strains confirmed whether the plantaricin operon exists in Lb. plantarum in a species- or strain-specific manner. While some Lb. plantarum strains possessed the plantaricin operon, not all strains possessed it (Fig. 1). This suggests that the plantaricin operon is strain-specific, not species-specific.

[0068] Example 2. Synthesis of plantaricin

[0069] The Lb. plantarumKM2 strain was confirmed to possess the bacteriocin genes PlnA (plantaricin A), PlnE (plantaricin E), PlnF (plantaricin F), PlnJ (plantaricin J), and PlnK (plantaricin K). The synthesized plantaricins were purified by HPLC, and the expected molecular weights were confirmed by MALDI-TOF-MS. All synthesized plantaricins were confirmed to be highly soluble in water (Table 1).

[0070] PlantaricinAmino acid sequenceSolubilityMS Found (Da)AAYSLQMGATAIKQVKKLFKKWGWWater2682.6EFNRGGYNFGKSVRHVVDAIGSVAGIRGILKSIRDMSO, water3546.2FVFHAYSARGVRNNYKSAVGPADWVISAVRGFIHGWater3702.8JGAWKNFWSSLRKGFYDGEAGRAIRRWater2929.4KRRSRKNGIGYAIGYAFGAVERAVLGGSRDYNKWater3502.7

[0071] Example 3. Antibacterial spectrum of synthesized plantaricin

[0072] The antibacterial activity of the synthesized plantaricins was evaluated using the disk diffusion method. To evaluate the antibacterial activity, four Gram-positive bacteria (B. cereus KCCM 11341, E. faecalis KCTC 2011, L. monocytogenes ATCC 19111, and S. aureus ATCC 12692) and five Gram-negative bacteria (A. xylosoxidans KCCM 40240, E. coli O157:H7 EDL 933, Flavobacterium sp. KCCM 11374, S. enterica KCCM 11862, and V. parahaemolyticus KCTC 2729; Table 2) were used as indicator strains. As a result, each synthetic plantaricin exhibited antibacterial activity against seven indicator strains except B. cereus and E. coli (Table 2). In particular, as a result of confirming the antibacterial activity in various combinations, the three combinations (spPlnAEJ, spPlnAEK, etc.) with added plantaricin A showed excellent activity against Flavobacterium sp. KCCM 11374 (Table 2). Considering that antibacterial activity was shown not only in the existing two combinations of peptide forms, EF and JK, but also in various combinations, it is judged that various combinations of plantaricin can show excellent antibacterial activity results depending on the target strain.

[0073] Seed strainKM2AEFJKEFEJEKJKFJFKAEAFAJAKAEFAEJAEKAJKAFJAFKGram-positive bacteriaBacillus cereusKCCM 11341++---------------------Enterococcus faecalisKCTC 2011++++-W+++++W++--+++W------Listeria monocytogenesATCC 19111-+++W+++++W++WWWWWW------Staphylococcus aureusATCC 12692+++--W-++-+W-+WW+-W--WWGram-negative bacteriaAlcaligenes xylosoxidansKCCM 40240++-W--------------W----Escherichia coliO157:H7EDL 933+++---------------------Flavobacteriumsp.KCCM 11374++++WW++++++++++++++-++++++-++++Salmonella entericaKCCM 11862+++WW---W--W--WWWW-W----Vibrio parahaemolyticusATCC 17802++W--W-++W-+W-++WW-WW-WW

[0074] Diameter of growth inhibition zone: -, 0.0 mm; W, 0.1-1.0 mm; +, 1.1-2.0 mm; ++, 2.1-3.0 mm; +++, >3.0 mm. Identical results from three experiments are presented.

[0075]

[0076] Example 4. Antibacterial activity of synthetic plantaricin against Flavobacterium sp. KCCM 11374

[0077] The antibacterial activity of synthetic plantaricins against Flavobacterium sp. KCCM 11374 was confirmed by the disk diffusion method and minimum inhibitory concentration (MIC) test. Synthetic plantaricins showed the highest antibacterial activity against Flavobacterium sp. KCCM 11374 (Table 2). Flavobacterium sp. is found in various environments such as dairy products, fish, and meat, and is a major cause of food spoilage. Among the synthetic plantaricin combinations against the Flavobacterium sp. KCCM 11374 strain, combinations containing spPlnA (Spontaneous plantaricin A), such as spPlnAEJ (Spontaneous plantaricin AEJ), spPlnAEK (Spontaneous plantaricin AEK), spPlnAFJ (Spontaneous plantaricin AFJ), and spPlnAFK (Spontaneous plantaricin AFK), showed excellent antibacterial activity (Table 2). The MIC results for spPlnAEJ (Spontaneous plantaricin AEJ), spPlnAEK (Spontaneous plantaricin AEK), spPlnAFJ (Spontaneous plantaricin AFJ), and spPlnAFK (Spontaneous plantaricin AFK) were 0.4, 0.4, 0.8, and 1.5 nmol / L, respectively. Therefore, spPlnAEJ (Spontaneous plantaricin AEJ) and spPlnAEK (Spontaneous plantaricin AEK) showed excellent results against Flavobacterium sp. in the disk diffusion method and minimum inhibitory concentration (MIC) test.

[0078] Example 5. Antibacterial activity of synthetic plantaricin against Flavobacterium sp. using Raman spectroscopy.

[0079] Raman spectroscopy is a technique that can rapidly measure intracellular components such as lipids, nucleic acids, and proteins by utilizing the characteristic of laser scattering by molecular resonance. Antibacterial activity was analyzed through quantitative evaluation of changes in physiological activity after treatment of Flavobacterium sp. with synthetic plantaricin. Flavobacterium sp. After treating KCCM 11374 with synthetic plantaricins spPlnA (Spontaneous plantaricin A), spPlnAEJ (Spontaneous plantaricin AEJ), spPlnAEK (Spontaneous plantaricin AEK), spPlnAFJ (Spontaneous plantaricin AFJ), and spPlnAFK (Spontaneous plantaricin AFK) at concentrations corresponding to half of the MIC value (0.1, 0.2, 0.2, 0.4, and 0.7 nmol / L, respectively), the spectral changes were confirmed using Raman spectroscopy. All spectral peaks showed an increasing pattern when treated with synthetic plantaricin (Fig. 2A). When treated with synthetic plantaricin, the peaks at 1001, 1133, 1289, 1457, 1535, 1658, 2731, 2931, and 3067 cm -1 The corresponding peaks increased, and each peak represents C=CH, CH deformation, CCC chain stretching, CC stretching, C≡C stretching, tryptophan, CH3-C aliphatic chain, amine vibration, and CCH aromatic ring stretching, indicating changes in the carbon and nitrogen components that constitute the cell wall (Fig. 2A).

[0080] To further investigate the difference in antibacterial activity of the synthetic plantaricin combinations, the Raman spectral range (400–1800 cm -1) was confirmed using PCA. The untreated control group not treated with synthetic plantaricin was located in the third quadrant, and the treatment group treated with synthetic plantaricin was distributed in the first, third, and fourth quadrants (Fig. 2B). In particular, spPlnA showed similar results to the untreated control group, but spPlnAEK, spPlnAEJ, and spPlnAFK showed distinct patterns from the untreated control group. In addition, the results of analysis using PLS-DA showed that the treatment group treated with synthetic plantaricin was different from the untreated control group (Fig. 2C). The treatment groups treated with synthetic plantaricins spPlnA, spPlnAEK, spPlnAEJ, and spPlnAFK were all located in the first, second, and third quadrants, whereas the untreated control group was located in the fourth quadrant, showing more distinct antibacterial activity (Fig. 2C).

[0081] Example 6. Cell lysis of Flavobacterium sp. KCCM 11374 by synthetic plantaricin

[0082] The effect of synthetic plantaricin treatment on the cell wall of Flavobacterium sp. KCCM 11374 was confirmed through transmission electron microscopy (TEM) analysis. The differences in cross-sections of Flavobacterium sp. KCCM 11374 after treatment with synthetic plantaricins spPlnA, spPlnAEJ, spPlnAEK, spPlnAFJ, and spPlnAFK were confirmed using TEM. The destruction of the cell wall of Flavobacterium sp. KCCM 11374 by synthetic plantaricin and the dissolution of cell wall fragments were confirmed. In particular, the dissolution of cell wall fragments was clearly confirmed in spPlnAEK, which showed high antibacterial activity (Fig. 3).

[0083] Flavobacterium sp. is a major cause of spoilage in fermented foods. Synthetic plantaricin has been shown to effectively inhibit spoilage by destroying the cell wall of Flavobacterium sp. and lysing the cells. Furthermore, plantaricin exhibits antibacterial activity against both Gram-positive and Gram-negative bacteria, demonstrating a broader spectrum of activity than existing natural antibacterial agents (e.g., nisin). These results suggest that plantaricin has the potential to be utilized as a promising natural antibacterial agent in the food industry.

[0084] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions merely represent preferred embodiments and are not intended to limit the scope of the present invention. In other words, the substantial scope of the present invention is defined by the appended claims and their equivalents.

[0085] Numerical ranges are inclusive of the values ​​defined in the ranges above. Any maximum numerical limitation given throughout this specification includes any lower numerical limitation, as if that lower numerical limitation were explicitly stated. Any minimum numerical limitation given throughout this specification includes any higher numerical limitation, as if that higher numerical limitation were explicitly stated. Any numerical limitation given throughout this specification will include any better numerical range within the broader numerical range, as if that narrower numerical limitation were explicitly stated.

[0086]

Claims

1. An antibacterial composition comprising, as an active ingredient, one or more plantaricin peptides derived from Lactiplantibacillus plantarum KM2 strain.

2. An antibacterial composition according to claim 1, characterized in that the Lactiplantibacillus plantarum KM2 strain is deposited under the accession number KCTC 14637BP.

3. An antibacterial composition according to claim 1, wherein the plantaricin peptide comprises an amino acid sequence selected from sequence numbers 1 to 5.

4. In the third paragraph, the antibacterial composition is characterized in that it is a combination of a plantaricin peptide comprising an amino acid sequence consisting of sequence number 1 and a plantaricin peptide comprising at least one amino acid sequence selected from sequence numbers 2 to 5.

5. In the first paragraph, the antibacterial composition is characterized in that it exhibits antibacterial activity against at least one strain selected from the group consisting of Flavobacterium sp., Enterococcus faecalis, Listeria monocytogenes, Staphylococcus aureus, Alcaligenes xylosoxidans, Salmonella enterica, and Vibrio parahaemolyticus.

6. An antibacterial composition according to claim 1, characterized in that the antibacterial composition is for oral administration.

7. A health functional food composition comprising an antibacterial composition selected from any one of claims 1 to 6 as an active ingredient.

8. A cosmetic composition comprising an antibacterial composition selected from any one of claims 1 to 6 as an active ingredient.

9. A food preservative composition comprising an antibacterial composition selected from any one of claims 1 to 6 as an active ingredient.

10. An oral therapeutic microbiome pharmaceutical composition comprising an antibacterial composition selected from any one of claims 1 to 6 as an active ingredient.

11. An oral microbiome regulating composition comprising an antibacterial composition selected from any one of claims 1 to 6 as an active ingredient.

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