Postbiotic with anti-corrosion function as well as preparation method and application thereof

Abstract

The invention discloses a metagen with a preservative function as well as a preparation method and application thereof, and belongs to the technical field of microbial fermentation. The metagen is prepared by the following method. The preparation method comprises the following steps: inoculating fermentation lactobacillus mucus Postbio-Q7 and lactobacillus paracasei Postbio-P6 into a fermentation culture medium, and carrying out fermentation; after the fermentation is finished, sterilizing the fermentation liquor to obtain post-prebiotics fermentation liquor; and then carrying out clarification and impurity removal treatment and concentration and desalination treatment on the metagen fermentation liquor, and freeze-drying to obtain the metagen. The metagen disclosed by the invention has an obvious inhibition effect on growth and development of putrefying bacteria such as pseudomonas putida and acinetobacter perforatum, so that the metagen has relatively good application potential and prospect in preservation of aquatic products.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbial fermentation, and in particular relates to a postbiotic with an antiseptic function, a preparation method and an application thereof. Background Art

[0002] Aquatic products are characterized by high protein, high quality fat and high nutrition. However, processed aquatic products are prone to spoilage during transportation and sales. Among them, microorganisms are the main cause of late deterioration. Their massive proliferation causes the decomposition of fat and protein in aquatic products, producing corrupt substances (alcohol, aldehyde, ketone, etc.) and odors (trimethylamine, hydrogen sulfide, etc.) that make them inedible. At present, the most commonly used technology for preserving aquatic products is 4°C low-temperature refrigeration and freezing. Most bacteria related to the spoilage of aquatic products, such as Shewanella, Pseudomonas, and Acinetobacter, are still active at low temperatures. They tend to form biofilms, attach to and live on aquatic products, causing aquatic food to spoil. Therefore, controlling the contamination of specific corrupt microorganisms in aquatic products is crucial to food safety.

[0003] In order to inhibit the growth and reproduction of spoilage bacteria, chemical preservatives are now widely used. However, the irrational use of chemical preservatives can easily cause human hypersensitivity reactions and chemical residues. At present, the use and dosage of chemical preservatives are strictly controlled. In recent years, natural biological preservatives produced by probiotic microorganisms and their secondary metabolites have gradually attracted people's attention due to their safety, stability and wide application. Studies have shown that some probiotics and metabolites can inhibit the growth of spoilage bacteria in aquatic products. Postbiotics are preparations of inanimate microorganisms and / or their components that are beneficial to the health of the host. They contain a variety of active ingredients, including bacterial components such as peptidoglycan and teichoic acid, as well as active metabolites such as vitamins, organic acids, bacteriocins, short-chain fatty acids, enzymes and amino acids. They have been shown to have antibacterial, anti-inflammatory, antioxidant and anti-cancer effects in in vivo and in vitro studies. Therefore, it is of certain application value to find a safe, effective and pollution-free antibacterial method to replace the use of traditional chemical preservatives. Summary of the invention

[0004] The present invention provides a method for preparing postbiotics with antiseptic function, comprising the following steps:

[0005] The fermentation Lactobacillus mucilaginosus Postbio-Q7 and Lactobacillus paracasei Postbio-P6 are inoculated into a fermentation medium for fermentation; after the fermentation is completed, the fermentation liquid is sterilized to obtain a postbiotic fermentation liquid; the postbiotic fermentation liquid is then clarified, impurity-removed, concentrated and desalted, and freeze-dried to obtain postbiotics.

[0006] In the above preparation method, the inoculation amount of the fermented mucus lactobacillus Postbio-Q7 and the Lactobacillus paracasei Postbio-P6 is selected from 3 to 7%, preferably 5%.

[0007] In the above preparation method, the fermentation conditions are selected from: fermentation temperature of 35-40°C, fermentation time of 36-48h, fermentation pH of 3.2-3.8; preferably: fermentation temperature of 37°C, fermentation time of 40h, fermentation pH of 3.5.

[0008] In the above preparation method, the formula of the fermentation medium is as follows: 20g glucose, 15g whey protein, 20g soy powder, 10g skim milk powder, 10g yeast powder, 0.5g Tween 80, 1g disodium hydrogen phosphate, 0.01g manganese sulfate, 0.1g magnesium sulfate, 0.02g leucine, 0.01g isoleucine, 1L distilled water, initial pH=7.0; before inoculation and fermentation, the fermentation medium needs to be sterilized at 105°C for 30min.

[0009] In the above preparation method, the sterilization conditions are selected from: pasteurization at 95°C for 40-100 minutes.

[0010] The present invention provides postbiotics prepared by the above method.

[0011] The present invention provides the use of the above-mentioned postbiotics in preparing a preparation for inhibiting spoilage bacteria; the spoilage bacteria are selected from Pseudomonas putida and / or Acinetobacter johnii.

[0012] The present invention provides the use of the above-mentioned postbiotics in food preservation; the food is preferably aquatic products.

[0013] The beneficial effects of the present invention are:

[0014] The postbiotics of the present invention have a significant inhibitory effect on the growth and development of spoilage bacteria such as Pseudomonas putida and Acinetobacter johnsonii, and thus have good application potential and prospects in the preservation of aquatic products. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 The inhibition zones of postbiotics against two spoilage bacteria; A is the Pseudomonas putida control group, B is the Pseudomonas putida experimental group, C is the Acinetobacter johnsonii control group, and D is the Acinetobacter johnsonii experimental group;

[0016] Figure 2 The ultrathin section microscope observation of postbiotics on two spoilage bacteria; A is the Pseudomonas putida control group, B is the Pseudomonas putida experimental group, C is the Acinetobacter johnsonii control group, and D is the Acinetobacter johnsonii experimental group;

[0017] Figure 3 The effect of postbiotics on the minimum inhibitory concentration of two spoilage bacteria;

[0018] Figure 4 The effect of postbiotics on biofilm formation of two spoilage bacteria strains;

[0019] Figure 5 The effect of postbiotics on DNA leakage of two spoilage bacteria;

[0020] Figure 6 The volcano plot of differentially expressed genes (left) and KEGG enrichment analysis of differentially expressed genes (right) of the transcriptome analysis of Pseudomonas putida by postbiotics;

[0021] Figure 7 The volcano plot of differentially expressed genes (left) and KEGG enrichment analysis of differentially expressed genes (right) of the transcriptome analysis of Acinetobacter johnsonii by postbiotics;

[0022] Figure 8 The effect of postbiotics on the color of aquatic products. DETAILED DESCRIPTION

[0023] The bacterial material used in the present invention is as follows:

[0024] The fermented mucus lactobacillus Postbio-Q7 and the paracasei Lactobacillus Postbio-P6 of the present invention are both known strains, which have been disclosed in Chinese patent CN 117305161 A (a postbiotic, its preparation method and application). Among them, fermented mucus lactobacillus (Limosilactobacillus fermentum) Postbio-Q7 and paracasei Lactobacillus (Lactobacillus paracasei) Postbio-P6 are both from the lactic acid bacteria strain bank of the Basic Research and Development Application Innovation Laboratory of Probiotics and Postbiotics of Qingdao Agricultural University, and are preserved in the General Microbiological Center of China Microbiological Culture Collection Administration Committee (referred to as CGMCC, address: No. 3, No. 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code 100101), and their preservation numbers are CGMCC No. 26238 and CGMCC No. 26237, respectively.

[0025] Other materials used in the present invention, if not otherwise stated, can be obtained through commercial channels. Other terms used in the present invention, unless otherwise specified, generally have the meanings commonly understood by those of ordinary skill in the art. The present invention is further described in detail below in conjunction with specific examples and with reference to data. The following examples are only for illustrating the present invention, and are not intended to limit the scope of the present invention in any way.

[0026] Example 1

[0027] To prepare postbiotics, the steps are as follows:

[0028] The fermented Lactobacillus mucilaginosus Postbio-Q7 and Lactobacillus paracasei Postbio-P6 strains were inoculated into MRS liquid culture medium, cultured at 37°C for 18 h, and subcultured twice to make the number of viable bacteria after activation reach 5-10×10 6 cfu / mL. The activated fermented mucus lactobacillus Postbio-Q7 and Lactobacillus paracasei Postbio-P6 were inoculated into the fermentation medium at an inoculation rate of 5%, and fermented at 37°C for 40 hours, with a fermentation pH of 3.5. After the fermentation, the fermentation broth was sterilized and pasteurized at 95°C for 70 minutes to obtain a postbiotic fermentation broth. The postbiotic fermentation broth was then clarified, impurity-removed, concentrated and desalted, and freeze-dried to obtain a postbiotic freeze-dried powder, i.e., the postbiotic described in the present invention.

[0029] The formula of the above fermentation medium is:

[0030] Glucose 20g, whey protein 15g, soy flour 20g, skim milk powder 10g, yeast powder 10g, Tween 80 0.5g, disodium hydrogen phosphate 1g, manganese sulfate 0.01g, magnesium sulfate 0.1g, leucine 0.02g, isoleucine 0.01g, distilled water 1L, initial pH = 7.0. Before inoculation and fermentation, the fermentation medium needs to be sterilized at 105°C for 30 minutes.

[0031] Example 2 Antibacterial Effect

[0032] 1. Preparation of bacterial suspension

[0033] The inhibitory effect of postbiotics on Pseudomonas putida (P. putida) and Acinetobacter johnsonii (A. johnsonii) was tested. Acinetobacter johnsonii and P. putida were isolated from grass carp fillets stored at low temperature (4°C) for 5 days and identified by Shanghai Saiheng Biotechnology Co., Ltd. (Shanghai, China). Acinetobacter johnsonii and P. putida were cultured in brain heart infusion (BHI) and nutrient broth (NB) medium (Qingdao Sun Biotechnology Co., Ltd.) at 30°C for 24h. Pure strains were obtained by centrifugation at 8000g for 10min and diluted to 10 with 0.9% sterile saline solution (SPSS, pH 7.4). 7 cfu / mL, for future use.

[0034] 2. Evaluation of antibacterial effect

[0035] First, place the Oxford cup on a sterile blank plate and add 700 μL of bacterial suspension (10 7CFU / mL) was added to an agar medium cooled to 37°C and shaken. After the medium was completely solidified, the Oxford cup was removed. 150 μL of postbiotic fermentation broth was added to the wells of the postbiotic group, and an equal volume of sterile saline was added to the other wells of the control group. The plate was transferred to 4°C and refrigerated for 4 hours, and then incubated at 30°C for 24 hours. Finally, the diameter of the inhibition zone was measured with a vernier caliper. Three replicates were performed for each sample, and the average value was taken as the final antibacterial effect.

[0036] The test results are as follows Figure 1 And as shown in Table 1.

[0037] Table 1 Diameters of inhibition zones of postbiotic fermentation broth against Pseudomonas putida and Acinetobacter johnsonii

[0038] strains Pseudomonas putida Acinetobacter johnsonii Inhibition zone size (mm) 30.85±0.85 25.32±0.66

[0039] Depend on Figure 1 As shown in Table 1, the diameters of the inhibition zones of the postbiotics against Pseudomonas putida and Acinetobacter johnsonii were 30.85±0.85mm and 25.32±0.66mm, respectively, both showing significant inhibitory effects. When the antibacterial diameter was greater than 20mm, it was determined that the strain was extremely sensitive to antimicrobial agents. In summary, the postbiotics had good antibacterial effects against Pseudomonas putida and Acinetobacter johnsonii.

[0040] 3. Ultrathin section microscopic observation of postbiotics on two spoilage bacteria

[0041] The bacterial suspension was transferred to a 2 mL sterile centrifuge tube and centrifuged at 4000 g for 15 min. 1.5 mL of postbiotic fermentation broth was added and incubated for 2 h as the postbiotic group, while an equal volume of sterile saline was added as the control group. After washing with sterile saline and centrifugation, the cells were fixed with 2.5% glutaraldehyde and refrigerated at 4 °C overnight. The bacteria were washed again and added to the dehydrating agent (acetone and ethanol) at a ratio of 1:1 (mass / volume), and repeated twice for 15 min each time. Thereafter, the dehydrated bacteria were embedded in 50% acetone and 50% EPON812 epoxy resin. The embedded blocks were transferred to 60 °C for 24 h polymerization. The sections were sliced ​​at 50-90 nm using an EM-UC6 / FC7 ultramicrotome (Leica Biosystems, Beijing, China), and the sections were removed and stained with 2% uranyl acetate and lead nitrate for 5 min. The samples were imaged using a Tecnai G2 Spirit electron microscope (FEI Corporation, Hillsboro, Oregon, US) at 90 kV.

[0042] The results are as follows Figure 2 As shown:

[0043] The bacterial bodies of the two bacterial control groups were observed to be plump, independent, and round, with no obvious damage. However, the postbiotic treatment caused obvious disorder and damage inside the bacterial bodies, and the bacterial organelles could not be identified. In addition, the surface of the bacterial bodies treated with postbiotics was damaged, causing the contents to flow out and causing adhesion between the bacterial bodies. The bacterial bodies became fragile and easily entangled, losing the original toughness of the cells.

[0044] 4. Effects of postbiotics on the minimum inhibitory concentration of two spoilage bacteria

[0045] The bacteria were inoculated into NB medium containing postbiotics (250, 125, 62.5, 31.25, 15.625, 7.8125, 3.9063, 1.9531, 0.9766, and 0.4883 μL / mL) at an inoculum size of 2% and incubated at 30°C for 24 h. The OD was measured by an ELISA instrument (Thermo Fisher Scientific, Qingdao, Shandong, China). 600 nm MIC was defined as the lowest concentration that inhibited 90% bacterial growth.

[0046] The results are as follows Figure 3 As shown:

[0047] As the concentration of postbiotics decreased, the colony increased significantly. The minimum inhibitory concentrations of Pseudomonas putida and Acinetobacter johnsonii after postbiotic treatment were 3.9063μL / mL and 15.625μL / mL, respectively. In the range of 250-15.625μL / mL, both strains were effectively inhibited. This shows that postbiotics exhibit strong antibacterial properties and can achieve inhibitory effects at low concentrations.

[0048] 5. Effects of postbiotics on biofilm formation of two spoilage bacteria strains

[0049] Dilute the bacteria in the logarithmic growth phase to 10 7 CFU / mL, the bacterial solution and culture medium were added to each well of a 96-well plate in a ratio of 1:1 (mass / volume). The blank control group used sterile saline instead of culture medium, and the experimental group used postbiotic fermentation broth instead. The bacteria were cultured at 30°C for 36 hours. The culture medium was removed by rinsing with sterile saline, and 200 μL of 1% crystal violet was added to each well for 15 minutes. After rinsing 3 times until the color became lighter, 200 μL of acetic acid was added to dissolve for 5 minutes. OD was measured using an enzyme reader 600nm value.

[0050] Biofilm inhibition rate calculation formula:

[0051]

[0052] Among them, OD C As the control group, ODP For the experimental group.

[0053] The test results are as follows Figure 4 As shown:

[0054] The biofilm inhibition rates of Pseudomonas putida and Acinetobacter johnsonii treated with postbiotics were 91% and 87%, respectively. The biofilm inhibition rate of the experimental group was significantly higher than that of the control group and the negative control group (p<0.05). The reason why the negative control group was lower than the control group may be that the osmotic pressure in the saline was too high, causing some bacteria to rupture. Postbiotics may decompose extracellular polysaccharides, thereby inhibiting the cell biofilm produced by bacteria.

[0055] 6. Effect of postbiotics on DNA leakage of two spoilage bacteria

[0056] 10 7 The bacterial solution with CFU / mL was centrifuged (8000g, 10min) to obtain bacterial sludge, and the above steps were repeated twice by washing with sterile saline. 1.5mL of postbiotic fermentation broth was added to the experimental group, and the same amount of sterile saline was added to the control group. The cells were incubated at 30℃ for 2h, and the OD was measured with an enzyme marker. 260 nm value.

[0057] DNA leakage rate calculation formula:

[0058]

[0059] Among them, OD C As the control group, OD P For the experimental group.

[0060] The test results are as follows Figure 5 As shown:

[0061] The DNA leakage rates of Pseudomonas putida and Acinetobacter johnsonii treated with postbiotic fermentation broth reached 90.7% and 80.2%, respectively. The DNA leakage rate in the experimental group was significantly different from that in the control group and the negative control group (p<0.05). The slightly higher DNA leakage rate in the negative control group may be related to cell rupture caused by excessive osmotic pressure, which is similar to the phenomenon in biofilms. In summary, postbiotic fermentation broth destroyed the bacterial cell membrane.

[0062] 7. Transcriptome analysis of Pseudomonas putida and Acinetobacter johnsonii by postbiotics

[0063] First, RNA was extracted using the RNeasy kit (Qiagen, Venlo, Netherlands). RNA integrity and total RNA were accurately measured using an Agilent 2100 bioanalyzer (Agilent Inc., Santa Clara, California, USA). The extracted RNA was removed from the total RNA by a probe to remove ribosomal RNA (rRNA) and enrich for mRNA. Libraries were established in a chain-specific manner and initially quantified using a Qubit 2.0 fluorometer (Thermo Fisher Science, Waltham, MA, USA). Finally, the amplified sequences were analyzed using an Illumina HiSeq 2500 platform (Beijing Biomarker Technology Co., Ltd., China).

[0064] According to data analysis, genes with |log2(FoldChange)|>=1&padj<=0.05 were selected as significantly different genes.

[0065] The test results are as follows Figure 6 and Figure 7 As shown:

[0066] like Figure 6 As shown in the middle left figure, compared with the Pseudomonas putida control group, there are a total of 2151 differentially expressed genes under postbiotic treatment, 1240 genes are significantly downregulated, and 911 genes are significantly upregulated. Figure 6 As shown in the middle right figure, after the postbiotic treatment, most genes of Pseudomonas putida were obviously involved in transmembrane transport, redox process, carbon metabolism, glycolysis metabolism, TCA cycle and ribosome metabolism. Figure 7 As shown in the middle left figure, compared with the control group of Acinetobacter johnsonii, there are a total of 1187 differentially expressed genes under postbiotic treatment, 610 genes are significantly downregulated, and 577 genes are significantly upregulated. Figure 7 As shown in the middle right figure, after postbiotic treatment, Acinetobacter Johannes was significantly involved in transmembrane transport, nucleotide biosynthesis, and glycolysis metabolism.

[0067] In terms of transmembrane transport, as shown in Table 2, compared with the Pseudomonas putida control group, the expression of PP4_RS23635, osmE, PP4_RS21210, kdpB and PP4_RS08775 in Pseudomonas putida treated with postbiotics was downregulated, while the expression of PP4_RS16945, icmH and PP4_RS14165 was upregulated. The PP4_RS23635 gene is related to efflux pump proteins, which may be a defense that enables cells to resist antibiotics and produce drug resistance by expelling harmful substances out of the cell. The osmE, PP4_RS21210 and kdpB genes are related to osmotic regulatory proteins, and the PP4_RS16945, icmH and PP4_RS14165 genes are related to membrane transport proteins. Postbiotics may inhibit the expression of osmotic regulatory proteins on the cell membrane, resulting in reduced transmembrane water transport and exchange. In order to maintain osmotic pressure, the expression of transport proteins is promoted. However, more metabolites infiltrate the cells, causing further damage and death.

[0068] Table 2 Significantly differentially expressed genes related to transmembrane transport in Pseudomonas putida

[0069]

[0070]

[0071] In addition, as shown in Table 3, compared with the control group of Acinetobacter johnii, the expression of A1_GM001112, A1_GM001283, A1_GM000881 and A1_GM002962 in the experimental group of Acinetobacter johnii was downregulated after postbiotic treatment, and the expression of A1_GM002020, A1_GM000240 and A1_GM002868 was upregulated. Among them, the A1_GM001283 and A1_GM001112 genes are associated with potassium transport ATPase and periplasmic binding protein, respectively. Studies have shown that potassium transport ATPase affects cellular nutrient exchange and transport. Periplasmic solute binding proteins act as molecular shuttles to help transport small solutes from the outer membrane of bacteria to the inner membrane. The A1_GM002020 gene is associated with efflux proteins, and IAA-lysine is secreted from protective cells. Postbiotics may inhibit the expression of transporters on the cell membrane, leading to their further infiltration and causing the expression of cellular stress.

[0072] Table 3 Significantly differentially expressed genes related to transmembrane transport in Acinetobacter johnsonii

[0073] Gene name description <![CDATA[log 2 FC]]> padj A1_GM001112 Plasma membrane-bound protein -3.4637 5.75E-20 A1_GM001283 Potassium transporting ATPase -2.3234 7.85E-38 A1_GM000881 Copper P-type ATPase -2.2281 1.83E-21 A1_GM002962 Zinc transporting ATPase -2.1677 2.70E-43 A1_GM002020 Efflux proteins 3.3174 2.30E-08 A1_GM000240 AAA ATPase domain 2.5627 1.47E-14 A1_GM002868 ABC transporter permeabilization protein 2.5287 4.28E-04

[0074] In terms of energy metabolism, as shown in Table 4, four genes of Pseudomonas putida are involved: glgA, PP4_RS03480, PF03971, and PP4_RS16775. glgA and PP4_RS03480 are related to glycogen synthesis and methylmalonate semialdehyde, respectively. Studies have found that glycogen synthase is indispensable for the storage of polysaccharides in Chlamydia. Coenzyme A acylated methylmalonate semialdehyde dehydrogenase is a member of the aldehyde dehydrogenase (ALDH) family and plays a vital role in the catabolic pathway of valine and thymine. The PF03971 and PP4_RS16775 genes are related to monomeric isocitrate dehydrogenase (rate-limiting enzyme) and chorismate mutase, respectively, which can inhibit mitochondrial energy production. Postbiotics may cause bacteria to lack energy by inhibiting the supply of energy metabolism, thereby further promoting cell apoptosis.

[0075] Table 4 Significantly differentially expressed genes related to energy metabolism of Pseudomonas putida

[0076]

[0077]

[0078] In addition, as shown in Table 5, the A1_GM003256 and KduI genes in the experimental group of Acinetobacter johnii are respectively related to shikimate dehydrogenase and 4-deoxy-L-threo-5-hexose uronic acid ketoisomerase (KduI protein). Studies have shown that shikimate dehydrogenase is an indispensable substance in the shikimate metabolic pathway of Escherichia coli. Studies have shown that kduI protein is used by Escherichia coli to decompose pectin. Vitamins may inhibit energy metabolism pathways, leading to cellular energy deficiency and apoptosis.

[0079] Table 5 Significantly differentially expressed genes related to energy metabolism of Acinetobacter johnsonii

[0080]

[0081] Four genes in the Pseudomonas putida experimental group were involved in DNA repair: PP4_RS15440, clpA, PP4_RS02920 and PP4_RS24530, especially PP4_RS15440 and PP4_RS24530 genes were more significant, see Table 6. PP4_RS15440 and PP4_RS24530 were related to ku protein and carboxypeptidase, respectively. Ku protein is involved in DNA repair function and several other DNA metabolic processes. Carboxypeptidase may promote DNA homologous recombination in Pseudomonas aeruginosa. Postbiotics may damage DNA structure by inhibiting the expression of DNA repair proteins, thereby leading to cell apoptosis.

[0082] Table 6 Significantly differentially expressed genes related to DNA repair in Pseudomonas putida

[0083] Gene name description <![CDATA[log 2 FC]]> padj PP4_RS15440 Ku protein -3.7538 8.80E-73 clpA ATP-dependent Clp protease -3.6195 4.73E-69 PP4_RS02920 α / β hydrolase 1.6374 6.80E-08 PP4_RS24530 DMT family transporters 1.5609 5.26E-08

[0084] In addition, the genes PadR, A1_GM002469, ArsR, A1_GM000973 and A1_GM002798 of the Acinetobacter johnsonii group are related to DNA damage and repair, as shown in Table 7. As factors controlling gene expression, the reduction of PadR and A1_GM002469 transcriptional regulators may lead to a reduction in mRNA, thereby affecting a series of cell growth and development metabolism. A1_GM000973 and A1_GM002798 are related to HTH transcriptional regulators. As factors controlling gene expression, their upregulation may lead to an increase in biofilms, thereby enhancing the cell's defense against adverse environments. Studies have shown that HTH regulators can be inhibited by targets. Postbiotics inhibit DNA-related regulatory factors, preventing cells from synthesizing proteins, resulting in weakened cell damage repair.

[0085] Table 7 Significantly differentially expressed genes related to DNA repair in Acinetobacter johnsonii

[0086]

[0087]

[0088] In summary, the postbiotics of the present invention destroy the transport proteins on the cell surfaces of Pseudomonas putida and Acinetobacter johnsonii, and further destroy the internal structure of the cells by infiltration, thereby affecting the energy metabolism of bacterial survival and damaging the DNA of the bacteria themselves, making it impossible to further repair and ultimately leading to bacterial apoptosis, thereby inhibiting the bacteria through the above-mentioned pathways.

[0089] 8. Effect of postbiotics on the color of aquatic products

[0090] Purchase fresh fish from a local supermarket and remove the head, scales and viscera. Cut the fish back muscle into pieces (3 × 2 × 0.5 cm 3 ), and then soaked it in postbiotic fermentation liquid and sterile water for 5 days.

[0091] The test results are as follows Figure 8 As shown:

[0092] On day 0, the fish meat of both groups was bright in color and elastic in texture. On day 5, the color of the fish meat without postbiotics gradually turned dark yellow, the freshness was poor, and the meat was soft and sticky, while the color of the fish meat soaked in postbiotics was close to that of day 0, proving that postbiotics can slow down the yellowing of fish meat and have a good antiseptic function.

[0093] 9. Effect of postbiotics on the total colony count of aquatic products

[0094] Chop the soaked fish meat into small pieces and transfer 3 g of the fish meat to a sterile bag. The ratio of fish meat to sterile saline is 1:9 (mass / volume). Homogenize the sample for 6 minutes. After homogenization, dilute the sample 10 times in a series. Apply the diluted sample to a plate, then incubate the plate at 37°C for 2 days and count the final total viable bacteria.

[0095] The test results are shown in Table 8:

[0096] The initial total colony count of fresh fish is usually 2-4log CFU / mL, and studies have shown that the maximum acceptable threshold for fish is 6log CFU / mL. Both the unsoaked postbiotic group and the soaked postbiotic group were below 2.3log CFU / mL on day 0. The unsoaked postbiotic group increased sharply to 7.12log CFU / mL on day 5, while the soaked postbiotic group only reached 5.48log CFU / mL on day 5, still remaining within the maximum acceptable threshold. There was a significant difference between the two groups (p<0.05). Therefore, postbiotics can effectively inhibit microbial growth and show effective antibacterial effects.

[0097] Table 8 Effect of postbiotic fermentation broth on the total colony count of aquatic products

[0098]

[0099] The above is only a preferred embodiment of the present invention, and does not limit the present invention in other forms. Any technician familiar with the profession may use the above disclosed technical content to change or modify it into an equivalent embodiment with equivalent changes. However, any simple modification, equivalent change and modification made to the above embodiment according to the technical essence of the present invention without departing from the technical solution of the present invention still belongs to the protection scope of the technical solution of the present invention.

Claims

1. A method for preparing a postbiotic with an antiseptic function, characterized in that: The steps include: The fermentation Lactobacillus mucilaginosus Postbio-Q7 and Lactobacillus paracasei Postbio-P6 are inoculated into a fermentation medium for fermentation; after the fermentation is completed, the fermentation liquid is sterilized to obtain a postbiotic fermentation liquid; the postbiotic fermentation liquid is then clarified, impurity-removed, concentrated and desalted, and freeze-dried to obtain postbiotics.

2. The preparation method according to claim 1, characterized in that: The inoculation amounts of the fermented mucus lactobacillus Postbio-Q7 and the paracasei Lactobacillus Postbio-P6 are both selected from 3 to 7%.

3. The preparation method according to claim 1, characterized in that: The fermentation conditions are selected from: fermentation temperature of 35-40°C, fermentation time of 36-48h, and fermentation pH of 3.2-3.

8.

4. The preparation method according to claim 1, characterized in that: The formula of the fermentation medium is as follows: 20g glucose, 15g whey protein, 20g soy flour, 10g skim milk powder, 10g yeast powder, 0.5g Tween 80, 1g disodium hydrogen phosphate, 0.01g manganese sulfate, 0.1g magnesium sulfate, 0.02g leucine, 0.01g isoleucine, and 1L distilled water; initial pH=7.

0.

5. The preparation method according to claim 1, characterized in that: The sterilization conditions are selected from: pasteurization at 95° C. for 40-100 min.

6. Postbiotics prepared by the method according to any one of claims 1 to 5.

7. Use of the postbiotic according to claim 6 in the preparation of a preparation for inhibiting spoilage bacteria.

8. The use according to claim 7, characterized in that: The spoilage bacteria are selected from Pseudomonas putida and / or Acinetobacter johnsonii.

9. Use of the postbiotics according to claim 6 in food preservation.

10. The use according to claim 9, characterized in that: The food is aquatic product.

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