Biological antibacterial agents and their applications

Abstract

The invention relates to the field of biopharmaceuticals, and discloses a biological antibacterial agent and its application, wherein the biological antibacterial agent contains biomass and auxiliary factors, wherein the biomass contains nisin, lysostaphin and lysozyme; the auxiliary factors are selected from at least one of dehydroacetic acid and its sodium salt, paraben and its sodium salt and organic acid salt; wherein, based on the total amount of the biological antibacterial agent, the content of nisin is 0.1-20% by weight, the content of lysozyme is 0.1-15% by weight, and the content of lysostaphin is 0.01-5% by weight. The antibacterial effect of the biological antibacterial agent provided by the invention is equivalent to or better than that of antibiotic products, the ethanol yield of the biological antibacterial agent provided by the invention is equivalent to or improved compared with that of antibiotics, and the biological antibacterial agent provided by the invention has significantly lower cost relative to antibiotics.

Description

Technical Field

[0001] The present invention relates to the field of bacteriostatic agents, and in particular to a biological bacteriostatic agent and application thereof. Background Art

[0002] In the 1940s, antibiotics were first used in animal husbandry and achieved good results, which were quickly promoted around the world. As a result, the negative effects and adverse reactions of antibiotic use have gradually been recognized by the world. Antibiotic resistance has become an increasingly serious threat to human health and has caused huge economic costs.

[0003] In recent years, the pace of banning antibiotics has gradually accelerated, and various countries have also introduced relevant policies to join the ranks of banning antibiotics. In 2015, my country successively issued the "Environmental Protection Law", "Food Safety Law" and "Feed Quality and Safety Management Standards". It is a general trend for the feed industry to achieve comprehensive antibiotic restriction and ban in the future.

[0004] Since low-temperature fermentation is prone to bacterial contamination during alcohol production, antibiotics such as Anjuntai and penicillin were used before. These antibiotics will enter the feed after ethanol distillation. With the development of antibiotic-free feed, the antibiotics used in the fermentation process will inevitably be replaced, and the development of a new type of biological antibacterial agent is imminent. There are also some biological antibacterial agents on the market, but they are weak in bactericidal ability and cannot meet the requirements for controlling miscellaneous bacteria during alcohol fermentation. Summary of the invention

[0005] In order to overcome the problems existing in the above-mentioned prior art, the present invention provides a biological antibacterial agent and its application. The biological antibacterial agent of the present invention has an antibacterial effect equivalent to or better than that of antibiotics, is friendly to the human body and the environment, and has low cost.

[0006] In order to achieve the above-mentioned object, the present invention provides a biological antibacterial agent on one hand, which contains biomass and auxiliary factors, wherein the biomass contains nisin, lysostaphin and lysozyme; the auxiliary factor is selected from at least one of dehydroacetic acid and its sodium salt, paraben and its sodium salt and organic acid salt;

[0007] Wherein, based on the total amount of the biological antibacterial agent, the content of nisin is 0.1-20% by weight, the content of lysozyme is 0.1-15% by weight, and the content of lysostaphin is 0.01-5% by weight.

[0008] The second aspect of the present invention provides the antibacterial application of the above-mentioned biological antibacterial agent.

[0009] The beneficial effects of the biological antibacterial agent provided by the present invention include:

[0010] The biological bacteriostatic agent provided by the present invention has an antibacterial effect equivalent to or better than that of antibiotics; the ethanol production of the biological bacteriostatic agent provided by the present invention is equivalent to or improved compared with that of antibiotics; the biological bacteriostatic agent provided by the present invention is friendly to the human body and the environment and is non-toxic compared with antibiotics; the cost of the biological bacteriostatic agent provided by the present invention is significantly lower than that of antibiotics. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1-9 The diagram is a graph showing the antibacterial effects of the biological antibacterial agents and antibiotics prepared in Examples 1-6 and Comparative Examples 1-3 on 47 strains of miscellaneous bacteria;

[0012] Fig.10 The biological antibacterial agent and antibiotic prepared in Examples 1-6 and Comparative Examples 1-3 are based on the actual materials of the factory, and 47 strains of miscellaneous bacteria are added to simulate the miscellaneous bacteria produced during the fermentation process. The alcohol content and acidity data of the samples after 72 hours of simulated bacterial fermentation are shown;

[0013] Fig.11 The biological antibacterial agent and antibiotic prepared in Examples 1-6 and Comparative Examples 1-3 use actual factory materials as raw materials, and additional 47 strains of miscellaneous bacteria are added to simulate the miscellaneous bacteria produced during the fermentation process. The residual sugar data of the sample after 72 hours of simulated bacterial fermentation is shown. DETAILED DESCRIPTION

[0014] The endpoints and any values ​​of the ranges disclosed in this article are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of each range, the endpoint values ​​of each range and the individual point values, and the individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this article.

[0015] In one aspect, the present invention provides a biological antibacterial agent, which contains biomass and auxiliary factors, wherein the biomass contains nisin, lysostaphin and lysozyme; the auxiliary factors are selected from at least one of dehydroacetic acid and its sodium salt, paraben and its sodium salt, and organic acid salt;

[0016] Wherein, based on the total amount of the biological antibacterial agent, the content of nisin is 0.1-20% by weight, the content of lysozyme is 0.1-15% by weight, and the content of lysostaphin is 0.01-5% by weight.

[0017] The present invention has a wide range of selection for the composition of biomass. In addition to nisin, lysozyme and lysostaphin, the biomass may also contain other biomass with antibacterial effects, all of which are within the protection scope of the present invention.

[0018] In the present invention, the term "nisin" has the conventional meaning in the art, and refers to a polypeptide substance produced by nisin, which is composed of 34 amino acid residues and has a molecular weight of about 3500 Da. The antibacterial effect of nisin is to interfere with the normal function of the cell membrane, causing cell membrane permeation, nutrient loss and membrane potential decrease, thereby leading to the death of pathogenic bacteria and spoilage bacteria cells, and can be obtained commercially.

[0019] According to the present invention, preferably, based on the total amount of the biological antibacterial agent, the content of nisin is 0.1-20% by weight, for example, it can be 0.1% by weight, 0.2% by weight, 0.3% by weight, 0.4% by weight, 0.5% by weight, 0.6% by weight, 0.7% by weight, 0.8% by weight, 0.9% by weight, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, 6% by weight, 7% by weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight, 16% by weight, 17% by weight, 18% by weight, 19% by weight, 20% by weight, and any value in the range formed by any two of these values; preferably 1-18% by weight, more preferably 5-15% by weight.

[0020] In the present invention, the term "lysozyme" has the conventional meaning in the art, and refers to an enzyme that can hydrolyze mucopolysaccharides in bacteria, also known as muramidase and N-acetyl muramid polysaccharide hydrolase, which can cleave the peptidoglycan structure of the bacterial cell wall, causing the bacteria to lyse and die, and it carries a large amount of positive charge in a near-neutral pH environment in the body, and can directly react with negatively charged viral proteins to inactivate the virus, and can be commercially available.

[0021] According to the present invention, preferably, based on the total amount of the biological antibacterial agent, the content of lysozyme is 0.1-15% by weight, for example, it can be 0.1% by weight, 0.2% by weight, 0.3% by weight, 0.4% by weight, 0.5% by weight, 0.6% by weight, 0.7% by weight, 0.8% by weight, 0.9% by weight, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, 6% by weight, 7% by weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight, and any value in the range formed by any two of these values; preferably 1-12% by weight, more preferably 3-10% by weight.

[0022] In the present invention, the term "lysostaphin" has the conventional meaning in the art, and refers to an enzyme that can hydrolyze the glycine peptide bonds in the peptidoglycan of bacterial cell walls to kill bacteria. It can be obtained commercially or homemade.

[0023] According to the present invention, preferably, based on the total amount of the biological antibacterial agent, the content of lysostaphin is 0.01-5% by weight, for example, it can be 0.01% by weight, 0.02% by weight, 0.03% by weight, 0.4% by weight, 0.05% by weight, 0.06% by weight, 0.07% by weight, 0.08% by weight, 0.09% by weight, 0.1% by weight, 0.2% by weight, 0.3% by weight, 0.4% by weight, 0.5% by weight, 0.6% by weight, 0.7% by weight, 0.8% by weight, 0.9% by weight, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, and any value in the range formed by any two of these values; preferably 0.1-3% by weight, more preferably 0.5-2% by weight.

[0024] According to the present invention, preferably, based on the total amount of the biostatic agent, the content of the biomass is 0.3-50% by weight, and the content of the auxiliary factor is 50-99.7% by weight.

[0025] More preferably, based on the total amount of the biostatic agent, the content of the biomass is 2-40% by weight, and the content of the auxiliary factor is 60-98% by weight.

[0026] Further preferably, based on the total amount of the biostatic agent, the content of the biomass is 9-35% by weight, and the content of the auxiliary factor is 65-91% by weight.

[0027] According to the present invention, preferably, the biomass further contains polylysine. This preferred embodiment can achieve a synergistic antibacterial effect and improve the antibacterial efficiency.

[0028] In the present invention, the term "polylysine" has the conventional meaning in the art, and refers to a cationic polymer of 25-30 lysine, which has a destructive effect on cell membranes, has a broad antibacterial spectrum, and can be obtained commercially.

[0029] According to the present invention, preferably, based on the total amount of the biological antibacterial agent, the content of polylysine is 0.1-8% by weight, preferably 1-7% by weight. This preferred embodiment is conducive to enhancing the antibacterial effect of the biological antibacterial agent.

[0030] According to the present invention, preferably, the dehydroacetic acid and its sodium salt are dehydroacetic acid and / or sodium dehydroacetate. With this preferred embodiment, the dehydroacetic acid and its sodium salt have a broad spectrum of antibacterial ability, can effectively penetrate into the cell body, inhibit the respiration of microorganisms, and further enhance the antibacterial effect of the biological antibacterial agent.

[0031] According to the present invention, preferably, the parabens and their sodium salts are selected from at least one of methyl paraben, ethyl paraben, propyl paraben, butyl paraben, methyl paraben sodium, ethyl paraben sodium, propyl paraben sodium and butyl paraben sodium. With this preferred embodiment, the parabens and their sodium salts have strong antibacterial properties, and can denature proteins in cells by destroying the cell membrane of microorganisms, and can inhibit the activity of respiratory enzymes and electron transfer enzymes of microbial cells, further enhancing the antibacterial effect of biological antibacterial agents. The present invention has a wide range of selection of parabens and their sodium salts, as long as the above purpose can be achieved. Methyl paraben is used in the embodiments of the present invention for exemplary description, but the present invention is not limited thereto.

[0032] The organic acid salt of the present invention cooperates with dehydroacetic acid and its sodium salt, paraben and its sodium salt to further improve the antibacterial property of biomass. According to the present invention, preferably, the organic acid salt is selected from at least two of sodium propionate, sodium gluconate and sodium diacetate. The organic acid salt can be a combination of any two enzymes, for example: sodium propionate and sodium gluconate, sodium diacetate and sodium propionate, sodium gluconate and sodium diacetate, etc.

[0033] Preferably, the organic acid salt is sodium propionate, sodium gluconate and sodium diacetate. This preferred embodiment is more conducive to improving the antibacterial performance of biomass.

[0034] Preferably, the weight ratio of sodium gluconate, sodium propionate and sodium diacetate is 1:0.4-3:0.5-5, more preferably 1:1-2:2-4.

[0035] Preferably, the auxiliary factor contains at least two of dehydroacetic acid and its sodium salt, paraben and its sodium salt and organic acid salt, and can be any combination of two of them. For example: dehydroacetic acid and its sodium salt and paraben and its sodium salt, dehydroacetic acid and its sodium salt and organic acid salt, and paraben and its sodium salt and organic acid salt.

[0036] Most preferably, the cofactor contains dehydroacetic acid and its sodium salt, paraben and its sodium salt, and an organic acid salt.

[0037] According to the present invention, preferably, based on the total amount of the biostatic agent, the content of dehydroacetic acid and its sodium salt is 10-90 weight%, for example, it can be 10 weight%, 15 weight%, 20 weight%, 25 weight%, 30 weight%, 35 weight%, 40 weight%, 45 weight%, 50 weight%, 55 weight%, 60 weight%, 65 weight%, 70 weight%, 75 weight%, 80 weight%, 85 weight%, 90 weight%, and any value in the range formed by any two of these values; preferably 30-80 weight%, more preferably 50-70 weight%.

[0038] According to the present invention, preferably, based on the total amount of the biostatic agent, the content of paraben and its sodium salt is 0.1-25% by weight, for example, it can be 0.1% by weight, 0.5% by weight, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, 6% by weight, 7% by weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight, 16% by weight, 17% by weight, 18% by weight, 19% by weight, 20% by weight, 21% by weight, 22% by weight, 23% by weight, 24% by weight, 25% by weight, and any value in the range formed by any two of these values; preferably 1-20% by weight, more preferably 3-15% by weight.

[0039] According to the present invention, preferably, based on the total amount of the biostatic agent, the content of the organic acid salt is 0.1-30% by weight, for example, it can be 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, and any value in the range formed by any two of these values; preferably 1-25% by weight, more preferably 5-15% by weight.

[0040] According to the present invention, preferably, based on the total amount of the biostatic agent, the content of dehydroacetic acid and its sodium salt is 10-90% by weight, the content of paraben and its sodium salt is 0.1-25% by weight, and the content of the organic acid salt is 0.1-30% by weight.

[0041] According to the present invention, more preferably, based on the total amount of the biostatic agent, the content of dehydroacetic acid and its sodium salt is 30-80% by weight, the content of paraben and its sodium salt is 1-20% by weight, and the content of the organic acid salt is 1-25% by weight.

[0042] According to the present invention, further preferably, based on the total amount of the biostatic agent, the content of dehydroacetic acid and its sodium salt is 50-70% by weight, the content of paraben and its sodium salt is 3-15% by weight, and the content of the organic acid salt is 5-15% by weight.

[0043] The present invention does not particularly limit the preparation method of the biological antibacterial agent, and the biological antibacterial agent can be prepared according to conventional methods, for example, by directly mixing the components according to predetermined amounts.

[0044] The second aspect of the present invention provides the antibacterial application of the above-mentioned biological antibacterial agent.

[0045] According to the present invention, preferably, the bacteria can be Gram-positive bacteria, Gram-negative bacteria, actinomycetes, molds, etc. For example, the bacteria are 47 strains of bacteria that can be screened out from the bacteria produced in the actual fermentation process of the factory.

[0046] The present invention will be described in detail below through examples.

[0047] Nisin and lysozyme were commercially available from Zhejiang Yinuo Biotechnology Co., Ltd., food grade;

[0048] Polylysine is a commercial product from Shandong Qilu Biotechnology Co., Ltd., food grade;

[0049] Sodium dehydroacetate is a commercial product of Shandong Shengxie Biotechnology Co., Ltd., food grade;

[0050] Methylparaben is a commercial product of Shengyuan Food Ingredients Co., Ltd., food grade;

[0051] Sodium gluconate is a commercial product of Shandong Baisheng Biotechnology Co., Ltd., food grade;

[0052] Sodium propionate is a commercial product of Shandong Hongguan Biotechnology Co., Ltd., food grade;

[0053] Sodium diacetate is a commercial product of Zhejiang Yinuo Biotechnology Co., Ltd., food grade;

[0054] The antibiotic product (Anjuntai) is a commercially available product of Liuzhou Longtai Technology Co., Ltd. and is industrial grade.

[0055] Examples 1-6 and Comparative Examples 1-3

[0056] The components in Table 1 were added in sequence and mixed evenly to prepare biological antibacterial agents, which were recorded as S1-S6 and D1-3.

[0057] Table 1

[0058]

[0059] Note: The values ​​in the table are weight %

[0060] The following test cases are all conducted in a sterile environment. In the following test cases, each parameter is carried out according to the alcohol intermediate product testing method used by the factory.

[0061] The optical density (OD) parameter was measured by an ultraviolet spectrophotometer at a wavelength of 600nm; the alcohol content parameter was measured with reference to the alcohol meter method in GB5009.225-2016.

[0062] The specific operation of acidity determination is as follows: After filtering the mature mash with absorbent cotton, take 1 mL of the filtrate into a 100 mL conical flask, add 10 mL of neutral distilled water (previously neutralized with 0.01 mol / L sodium hydroxide standard titration solution using phenolphthalein as an indicator) and 1 drop of phenolphthalein indicator, and titrate with 0.1 mol / L sodium hydroxide standard solution until it turns slightly red. The end point is when it does not fade within 30 seconds. Acidity = volume of sodium hydroxide standard solution consumed in titration × concentration of sodium hydroxide standard solution × 10 / (volume of sample taken × 0.1).

[0063] Determination of volatile acid: Accurately measure 50mL of distilled liquor, pour into a conical flask, add 2 drops of phenolphthalein indicator, and titrate with 0.1mol / L sodium hydroxide solution until it turns light pink. The result is calculated as 1mL of 0.1mol / L sodium hydroxide standard titration solution consumed by 10mL of sample as one acidity unit. Volatile acid = volume of sodium hydroxide standard solution consumed in titration × concentration of sodium hydroxide standard solution × 10 / (volume of sample × 0.1).

[0064] Residual reducing sugar: Take 10g of saccharified mash, pour it into a 250mL conical flask, add water to make up to 250mL, mix well, filter with absorbent cotton, take 5mL of the filtrate and calculate sugar according to Fehling's method. Residual reducing sugar = (blank consumed glucose standard solution volume - sample consumed glucose solution volume) × glucose standard solution concentration × 100 / (sample volume × 5 / 250).

[0065] Residual total sugar: Measure 50mL of fermented mash and place it in a 250mL conical flask, add 40mL of water, add 10mL of 20% hydrochloric acid, plug the bottle mouth with a rubber stopper of a 1m long glass tube, convert in a boiling water bath for 60min, take out and cool, neutralize with 20% sodium hydroxide to slightly acidic and transfer to a 250mL volumetric flask and dilute to the scale, shake well and filter with absorbent cotton. Take 10mL of the filtrate and place it in a conical flask containing 5mL of Fehling's reagent A and B and 20mL of water, titrate with 0.25% glucose standard solution, and the following determination is the same as reducing sugar, and a blank test is performed at the same time. Residual total sugar = (blank titration consumed glucose solution volume - sample titration consumed glucose volume) × glucose solution concentration × 100 / (absorbed sample volume × 10 / 250).

[0066] Fehling's method: Take 5 mL of Fehling's solution A and solution B, put them into a 250 mL conical flask, add 30 mL of water, mix well, and heat on an electric stove with asbestos wire mesh (boil within 2-3 minutes). After boiling, use a burette to pre-add less than 0.5-1 mL of the sample in the preliminary experiment, place it on the electric stove and heat it to boiling. Keep boiling for 2 minutes, then drop 2 drops (0.1 mL) of 0.5% methylene blue indicator, and then drop glucose solution at a rate of 2-3 seconds / drop until the blue color of the test solution disappears and begins to turn red, which is the end point.

[0067] Test Example 1:

[0068] This test case is a single-bacteria antibacterial experiment on 47 strains of bacteria that can be screened out from the bacteria produced in the actual fermentation process.

[0069] 1) Preparation of indicator bacteria solution:

[0070] 47 strains of bacteria were cultured in YPD medium (2 wt% glucose, 1 wt% yeast, 2 wt% peptone and 95 wt% distilled water) for about 18 h to obtain bacterial liquid; the OD value of each bacterial liquid was diluted to 1, and then the bacterial liquid was diluted 1000 times (980uL YPD + 20uL bacterial liquid) to obtain the indicator bacterial liquid.

[0071] 2) Preparation of acidic YPD medium (simulating the environment of the fermentation tank):

[0072] Take 80 mL YPD, adjust the pH to 4.2 with H2SO4, sterilize with saturated steam at 121°C, and then add acetic acid and lactic acid to a final concentration of 1 g / L.

[0073] 3) Antibacterial experiment:

[0074] Experimental group: Take 80mL of acidic YPD medium, add the biological antibacterial agents prepared in Examples 1-5 and Comparative Examples 1-4 to a final concentration of 5mg / L, mix well, and obtain an antibacterial culture solution. Pipette 950uL of the antibacterial culture solution and add it to wells 1-47 of a 96-well plate, and add 50uL of the indicator bacterial solution of 47 strains of miscellaneous bacteria to wells 1-47 to obtain a sample;

[0075] Blank control: 950uL of acidic YPD medium was added to wells 1-47 of a 96-deep-well plate, and 50uL of indicator bacterial solution of 47 strains of bacteria was added to wells 1-47 to obtain samples;

[0076] Control group 1: Compared with the experimental group, the difference is that no biostatic agent was added, and antibiotics were added to a final concentration of 5 mg / L;

[0077] All samples were placed in a microporous shaker and cultured at 32°C. 100uL of sample was taken at 0 and 24h to measure OD 600The inhibition rate was calculated with control group 1 as reference. The results are as follows Figure 1-9 shown.

[0078] The single-bacteria antibacterial experiment can characterize the types and quantities of bacteria that the antibacterial agent can inhibit. From the results of the antibacterial rate, the antibacterial rates of Examples 1-3 for 47 strains of bacteria are higher than or equal to those of the antibiotic product ( Figure 1-3 ), the antibacterial rates of Examples 4-6 for most of the bacteria were higher than or equal to those of the antibiotic products, and the antibacterial rates for a small number of bacteria (such as strain numbers 19, 29, 30, etc.) were slightly lower than those of the antibiotic products ( Figure 4-6 ), the antibacterial rates of comparative examples 1-3 for most of the miscellaneous bacteria were significantly lower than those of antibiotic products ( Figure 7-9 ).

[0079] It can be seen from the results of Test Example 1 that the biological antibacterial agent of the present invention can effectively inhibit single bacteria, and its antibacterial effect is equivalent to or better than that of antibiotics.

[0080] Test Example 2:

[0081] This test case is a simulated bacterial fermentation experiment using actual factory materials as raw materials and adding 47 strains of miscellaneous bacteria.

[0082] Step 1: Take 200 mL of liquefied mash raw material from the factory, add 47 strains of bacteria described in Test Example 1, and add each strain of bacteria to a final concentration of 10 6 / mL, cool to 32°C, and adjust pH to 4.2-4.4 with sulfuric acid;

[0083] Step 2: Add 0.1 g Angel Super Wine Yeast Dry Powder, 0.041 g saccharifying enzyme and 0.002 g acid protease to each bottle of liquefied mash for subsequent synchronous saccharification and fermentation;

[0084] Step 3, three parallel experiments were carried out in each group, 5 mg / L of antibiotics were added to the control group, and 5 mg / L of the final concentration of the biological antibacterial agent prepared in Examples 1-5 and Comparative Examples 1-4 were added to the experimental group, mixed at a speed of 100 r / min, the fermentation temperature was 32° C., and the fermentation time was 72 h;

[0085] Step 4: After fermentation is completed, 100 mL of the mature mash sample is mixed with 100 mL of deionized water, and then distilled to determine the alcohol content;

[0086] Step 5: Take the remaining sample to determine the residual sugar, acidity and volatile acid.

[0087] Alcohol content is used to characterize the effect of adding different types of antibacterial agents on the fermentation effect of Angel Super Wine Yeast. Fig.10), the alcohol content of Examples 1-3 can reach or exceed that of antibiotic products. The alcohol content of Examples 4-5 is slightly lower than that of antibiotic products, and the alcohol content of Example 6 and Comparative Examples 1-3 is significantly lower than that of antibiotic products.

[0088] The determination of acidity and volatile acidity is used to characterize the inhibitory effect of antibacterial agents on miscellaneous bacteria. Fig.10 ), the acidity and volatile acidity of Examples 1-3 were lower than or equal to the antibiotic product, the acidity and volatile acidity of Examples 4-5 were slightly higher than the antibiotic product group, and Example 6 and Comparative Examples 1-3 were significantly higher than the antibiotic product group.

[0089] This shows that the biological antibacterial agents of Examples 1-3 are more conducive to the fermentation of super wine yeast while achieving a better antibacterial effect than the antibiotic products; the biological antibacterial agents of Examples 4-5 are slightly weaker than the antibiotic products, but basically do not affect the normal fermentation of super wine yeast; the biological antibacterial agents of Example 6 and Comparative Examples 1-3 have a certain inhibitory effect on the fermentation of super wine yeast compared with the antibiotic products.

[0090] Residual reducing sugar and residual total sugar are used to characterize the thoroughness of fermentation. Fig.11 ), the residual reducing sugar and residual total sugar of Examples 1-3 are lower than those of the antibiotic product, the reducing sugar and residual total sugar content of Examples 4-5 are slightly higher than those of the antibiotic product, and those of Example 6 and Comparative Examples 1-3 are significantly higher than those of the antibiotic product group. This shows that the biological antibacterial agents of Examples 1-3 are more conducive to the utilization and fermentation of sugar by super-wine yeast than the antibiotic product; the biological antibacterial agents of Examples 4-5 are slightly weaker than those of the antibiotic product in terms of the effect of super-wine yeast fermentation of sugar; and the biological antibacterial agents of Example 6 and Comparative Examples 1-3 are significantly worse than those of the antibiotic product.

[0091] At the same time, the use costs of the antibiotic products and the biological antibacterial agents prepared in Examples 1-6 are calculated, as shown in Table 2. As can be seen from Table 2, the use costs of the formulas corresponding to Examples 1-6 are all lower than those of the antibiotic products. If applied to fuel ethanol production enterprises, on the one hand, it meets the demand for antibiotic-free replacement, and on the other hand, it is beneficial to fuel ethanol enterprises to reduce costs and increase efficiency.

[0092] Table 2

[0093]

[0094]

[0095] Note: The usage cost is calculated based on the average ethanol content of 110g / L obtained by fermentation.

[0096] It can be seen from the results of Test Example 2 that, compared with antibiotics, the biological antibacterial agent of the present invention can better inhibit the strains produced in the actual material fermentation process. It can be seen that the biological antibacterial agent of the present invention can partially or completely replace antibiotics, and reduce the use cost of the antibacterial agent without affecting the ethanol production.

[0097] The preferred embodiments of the present invention are described in detail above, but the present invention is not limited thereto. Within the technical concept of the present invention, the technical solution of the present invention can be subjected to a variety of simple modifications, including the combination of various technical features in any other suitable manner, and these simple modifications and combinations should also be regarded as the contents disclosed by the present invention and belong to the protection scope of the present invention.

Claims

1. A biological antibacterial agent, characterized in that: The biological antibacterial agent contains biomass and auxiliary factors, wherein the biomass contains nisin, lysostaphin and lysozyme; the auxiliary factors are composed of dehydroacetic acid and its sodium salt, paraben and its sodium salt, and organic acid salt; Wherein, based on the total amount of the biological antibacterial agent, the content of nisin is 1-18% by weight, the content of lysozyme is 1-12% by weight, the content of lysostaphin is 1-3% by weight; the content of dehydroacetic acid and its sodium salt is 30-80% by weight, the content of paraben and its sodium salt is 1-18% by weight, and the content of organic acid salt is 3-18% by weight; Wherein, the organic acid salt consists of sodium propionate, sodium gluconate and sodium diacetate.

2. The bio-bacteriostatic agent according to claim 1, wherein Based on the total amount of the biostatic agent, the content of the biomass is 2-40% by weight, and the content of the auxiliary factor is 60-98% by weight.

3. The biological antibacterial agent according to claim 1 or 2, wherein Based on the total amount of the biological antibacterial agent, the content of the biomass is 9-35% by weight, and the content of the auxiliary factor is 65-91% by weight.

4. The biostatic agent according to claim 3, wherein: Based on the total amount of the biological antibacterial agent, the content of nisin is 5-15% by weight, the content of lysozyme is 3-10% by weight, and the content of lysostaphin is 1-2% by weight.

5. The biological antibacterial agent according to claim 1, wherein The biomass also contains polylysine.

6. The biostatic agent according to claim 5, wherein: Based on the total amount of the biostatic agent, the content of polylysine is 0.1-8% by weight.

7. The biological antibacterial agent according to claim 6, wherein Based on the total amount of the biostatic agent, the content of polylysine is 1-7% by weight.

8. The biological antibacterial agent according to claim 1, wherein The dehydroacetic acid and its sodium salt are dehydroacetic acid and / or sodium dehydroacetate.

9. The biological antibacterial agent according to claim 1, wherein The paraben and its sodium salt are selected from at least one of methyl paraben, ethyl paraben, propyl paraben, butyl paraben, methyl paraben sodium, ethyl paraben sodium, propyl paraben sodium and butyl paraben sodium.

10. The biostatic agent according to claim 1, wherein: Based on the total amount of the bio-bacteriostatic agent, the content of dehydroacetic acid and its sodium salt is 50-70% by weight, the content of paraben and its sodium salt is 3-15% by weight, and the content of organic acid salt is 5-15% by weight.

11. Antibacterial application of the biological antibacterial agent according to any one of claims 1 to 10 in alcohol fermentation.

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