CN107267410B

CN107267410B - Antibacterial method

Info

Publication number: CN107267410B
Application number: CN201710321477.3A
Authority: CN
Inventors: 姚圣; 阪本忠幸
Original assignee: Shanghai Jiuzhou Environmental Protection Technology Co ltd
Current assignee: Shanghai Jiuzhou Environmental Protection Technology Co ltd
Priority date: 2017-05-09
Filing date: 2017-05-09
Publication date: 2021-01-26
Anticipated expiration: 2037-05-09
Also published as: CN107267410A

Abstract

The invention belongs to the field of microorganisms, and particularly relates to an antibacterial method using a compound microbial agent. The effective components of the selected microbial agent are 1-5 parts by weight of spherical lysine bacillus and 1-5 parts by weight of bacillus thuringiensis. The invention provides a method for killing specific bacteria on an object without directly contacting the object with an antibacterial agent during use.

Description

Antibacterial method

Technical Field

The invention belongs to the field of microorganisms, and particularly relates to an antibacterial method of a compound microbial agent.

Background

With the continuous development of the living standard of people, the waste water and waste generated in daily production and life of residents are increased gradually, so that the continuously diffused malodorous gas always exists in farms, garbage piles and polluted water bodies, and the malodorous gas directly influences the living quality of people around, and even harms the health of people. In addition, with the pursuit of the quality of indoor life by residents,

more and more appliances in the toilet such as a bathtub, a toilet bowl, a urinal, a hand sink, a storage rack, a washing machine and the like are easy to generate daily cleaning dead corners, the toilet is inconvenient to clean completely, wastes time and labor, dirt is generated in the toilet after a long time, the peculiar smell in the toilet influences the daily life of people, pests are bred, and the living quality is seriously influenced.

The traditional method for preventing mildew and eliminating peculiar smell usually uses essence to cover peculiar smell or uses a physical air exhaust mode; or the ozone oxidizes and decomposes the peculiar smell; or by chemical means. Although the methods can also alleviate the peculiar smell to a certain degree, the method can not solve the problem fundamentally and really, in addition, the essence and the chemical deodorization or antibacterial method have the possibility of secondary pollution, if the method is not used properly, even harmful to human bodies, for example, the common use of concentrated hydrochloric acid and 84 disinfectant leads to casualty accidents.

The kitchen is used as a food processing space, oil smoke is attached to all parts of the kitchen when dishes are made, water consumption in the kitchen is large and wet, mold breeding is easily caused, the wall surface of the kitchen is discolored after a long time, and pests are bred.

In recent years, the microbial technology has attracted people's attention to the treatment of environmental pollution, and its advantages of high efficiency, economy and no secondary pollution make it more worthy of popularization than physical and chemical methods. The fields of deodorization by using microorganisms, fruit and vegetable cleaning, aquaculture, mildew prevention and the like are more and more accepted by people.

At present, chemical cleaning agents and frequent cleaning are mostly adopted in the prior art for deodorizing the toilet to achieve the aim of keeping the toilet free of peculiar smell. Chinese patent 104946544A discloses a microbial agent for deodorizing municipal domestic waste, public toilets and livestock farms, which comprises Candida utilis, Lactobacillus plantarum, Streptococcus lactis, Saccharomyces cerevisiae, Streptococcus thermophilus, Bacillus licheniformis and Bacillus natto.

Beneficial microorganisms contained in the deodorizing composite microbial agent can convert putrefaction degradation of putrefactive organisms in refuse dumps, public toilets and livestock farms into fermentation decomposition, absorb and degrade generated malodorous gases such as hydrogen sulfide and ammonia gas, eliminate the malodorous of garbage and garbage leachate, and simultaneously beneficial bacteria such as phytobacter can generate lactic acid to inhibit the growth of putrefactive bacteria, so that the generation of malodorous gases is fundamentally solved, and a garbage stacking environment capable of eliminating the malodorous gases and preventing diseases from spreading is formed.

However, the microbial inoculum contains beer yeast, alcohol and carbon dioxide are produced in an anaerobic environment, and the carbon dioxide can neutralize lactic acid produced by lactobacillus plantarum, streptococcus lactis and streptococcus thermophilus in water, namely, the using environment cannot be acidified. The environment adaptability is poor, and the deodorization effect is weak.

At present, basically all kitchen and bath cleaners in the market need to be in contact with an object to be cleaned so as to achieve a sterilization effect. Common kitchen and bath cleaning methods include wiping with a wipe soaked with an antimicrobial solution, or applying the antimicrobial directly to the target, or spraying the antimicrobial solution onto the target in a spray. However, such a method of bringing the antibacterial agent into direct contact with the object requires that the object be moved or that a sufficient space be left around the portion of the object to be cleaned, and thus greatly restricts the object and the portion that can be cleaned. And the common chemical cleaning agent has larger irritation to the human body when in use, thereby causing secondary damage to the human body.

Disclosure of Invention

The present inventors have conducted intensive studies to solve the above problems and, in combination with the prior art, have found that some common antibacterial strains have a bactericidal effect and exhibit excellent antibacterial activity without contacting an antibacterial target at the time of use, thereby completing the present invention.

An antibacterial method is characterized in that under the condition that an antibacterial agent is used in a closed space and is not in contact with a target object, the antibacterial agent inhibits the growth of specific bacteria;

the effective components of the antibacterial agent comprise 1-5 parts by weight of spherical lysine bacillus and 1-5 parts by weight of bacillus thuringiensis;

the specific bacteria are one or more than two of cladosporium, cladosporium sphaerothecoides, alternaria alternata, curvularia and rhizoctonia solani;

the volume of the closed space is less than 10L.

Furthermore, the antibacterial agent also comprises 250 parts by weight of microzyme and 600 parts by weight of photosynthetic bacteria; the preferable composition of the microbial inoculum comprises 2 weight parts of spherical lysine bacillus, 2 weight parts of bacillus thuringiensis, 200 weight parts of yeast and 500 weight parts of photosynthetic bacteria.

Further, the yeast is Angel yeast; the photosynthetic bacteria are eel photosynthetic bacteria.

Furthermore, the antibacterial agent also contains poultry manure treated at a high temperature of 60-150 ℃; the mass ratio of all the microbial bacteria in the microbial inoculum to the culture medium is 1-10: 100

Further, the poultry manure is selected from at least one of cow manure, pig manure and chicken manure.

Use of the antibacterial method according to any one of the preceding claims for mould proofing and disease prevention in plants.

The preparation method of the antibacterial agent comprises the following steps:

(1) preparing a culture medium suitable for culturing the lysine bacillus sphaericus, the bacillus thuringiensis, the yeast and the photosynthetic bacteria according to the selected strain; (2) and adding selected amounts of the lysine bacillus sphaericus, the bacillus thuringiensis, the yeast and the photosynthetic bacteria into the culture medium, and culturing at 36-38 ℃ to obtain the composite microbial deodorant and mildewproof agent.

The specific preparation steps of the culture medium comprise: the poultry manure powder is taken and added with water under the temperature of 30-50 ℃ until the humidity is 20-50 percent, and the poultry manure powder is obtained by cultivation.

In the step (2), the method further comprises the step of activating the selected strain in a proper activating solution.

The activation solution of the activation step is the same as the medium.

The invention has the following advantages:

1. the method is applied to the field of polluted water, pipelines and textiles, can perform deodorization and mildew prevention under the condition that a microbial inoculum is not contacted with a target object in use, and is suitable for narrow spaces.

The special microbial flora capable of converting or degrading the malodorous substances is used for decomposing, absorbing and metabolizing the malodorous gases such as hydrogen sulfide, sulfur dioxide, ammonia, nitrogen, phosphorus, carbon disulfide, methane and the like, harmful substances, pathogenic bacteria and the like emitted by water bodies, sewer pipelines and the like to purify and decompose the malodorous gases, the harmful substances, the pathogenic bacteria and the like, so that the malodorous gases, the harmful substances, the pathogenic bacteria and the like can be converted into harmless and odorless substances, the pollution of the malodorous gases, the harmful substances, the pathogenic bacteria and the like to the air is reduced, and. The purposes of thoroughly improving water quality, purifying environment and protecting human health are achieved.

2. In the preparation process of the composite microbial agent for deodorizing and preventing mold adhesion, poultry manure is used as a culture medium, and all preparation processes are carried out under a solid condition, so that the operation step of preparing powder from liquid through drying is omitted. The cost is reduced, and the preparation process is more efficient.

3. The present invention utilizes photosynthetic bacteria that can grow and metabolize in a completely inorganic environment that can sequester inorganic contaminants such as H₂S、NH₃The substances are converted into sulfur-containing proteins and become a component of novacells, or further converted into NO with low toxicity₃ ⁺And molecular sulfur. Meanwhile, photosynthetic bacteria can also reduce CO in the air by utilizing photosynthesis₂Thereby improving the quality of the air. The photosynthetic bacteria can quickly and effectively oxidize the odorous substances such as ammonia, hydrogen sulfide and the like into odorless, nontoxic and nonvolatile nitrate, sulfur-containing protein and microbial cytoplasm in the water body. Therefore, the odor released from the nitrogen-and sulfur-containing substances is more likely to be generatedIs effectively controlled. And the environmental adaptability and strong deodorization and pathogenic bacteria inhibition capability of the bacillus sphaericus are utilized. A large number of beneficial microorganisms and produced decomposition enzymes thereof can decompose organic matters such as ammonia nitrogen, hydrogen sulfide, mercaptan, methane, phosphorus and the like produced in a water body or a sewer pipeline (including a primary pollution source), inhibit harmful microorganisms, carry out continuous beneficial fungus reproduction, degrade pollutants, deodorize, purify polluted water and reduce dirt. The combined action of various microorganisms is more beneficial to absorbing and decomposing harmful gases with foul smell in water bodies and sewer pipelines.

4. The microbial inoculum used in the composite microbial deodorization and mildew-proof method is powder, can be used independently or mixed with carriers such as liquid, paste, absorbent gel and the like, and has various and simple use types. The microorganisms in the microbial inoculum can also produce the effects of antibiosis and poison suppression. No secondary pollution and secondary disaster are generated, the original ecological environment is restored, and the self-cleaning capability is improved.

Drawings

FIG. 1 is a schematic view showing the placement of a culture dish in example 1.

FIG. 2 is a schematic view showing the placement of a 1.3L vessel culture dish in example 2.

FIG. 3 is a schematic diagram showing the placement of a 10L vessel culture dish in example 3.

FIG. 4 is a schematic diagram of the experimental effect of example 1.

Detailed Description

The spherical lysine bacillus adopted by the invention is purchased from CGMCC (number: 1.270), and the Bacillus thuringiensis is purchased from CGMCC (number: 1.785); the yeast is Angel yeast, and the photosynthetic bacteria is Monopteri albi photosynthetic bacteria. The selected experimental strain is purchased from CGMCC.

The preparation method of the antibacterial agent used in the examples is as follows:

an antibacterial agent A:

1. in a sterile production workshop, the temperature control system was turned on with the 150kg tank as standard and the culture tank was allowed to rise to 37 ℃. (the temperature of the temperature control system is controlled at 37 ℃).

2. 100kg of chicken manure sterilized at 100 ℃ in a high-temperature sterilizer was added to the culture tank. Opening a stirring system of the fermentation tank to stir for 10 minutes; obtaining the culture medium.

3. Opening the discharge port of the fermentation tank to discharge 1kg of culture medium, mixing with 2kg of water to obtain an activation solution for activating strains, and adding 2000g of Angel yeast powder into 1.2kg of the activation solution. 3000g of eel photosynthetic bacteria powder is added into 1.2kg of the activation solution. 20g of spherical lysine bacillus and 20g of bacillus thuringiensis are respectively and uniformly mixed with 300g of activating solution. This step is used to activate the bacterial species.

4. The activated saccharomycetes, photosynthetic bacteria, spherical lysine bacillus and bacillus thuringiensis are sequentially added into a culture tank for fermentation, and water is added until the humidity in the culture tank is 25%.

5. And starting a stirring system to stir for 15 minutes, starting a temperature control system, and controlling the temperature to be 37 ℃ to culture for 7 days to finish production. (Note: the stirring system was started in the morning, in the middle of the day, and in the evening for 15 minutes each day for 7 days).

6. And (3) detection: the PH value is between 6 and 7, and the number of the beneficial viable bacteria is more than 5 hundred million per gram.

7. And packaging by using packaging equipment.

Antibacterial agent B

The differences from the preparation of the antibacterial agent A are 1500g of yeast, 4000g of photosynthetic bacteria, 10g of spherical lysine bacillus and 10g of bacillus thuringiensis.

In the antibacterial experiments of the examples, the selected experimental strains were purchased from CGMCC and shown in the following table

TABLE 1 Experimental strains used in the examples

The media used were:

1. nutrient agar medium (NA): 5.0g/L meat extract, 10.0g/L soybean meal, 5.0g/L sodium chloride and 15.0g/L agar. The pH value is 7.0 +/-0.1.

2. Potato dextrose agar medium (PDA): 200.0g/L of potato, 20.0g/L of glucose and 15.0g/L of agar.

Example 1 antimicrobial Activity test in a Small space

The experiment is divided into three groups, namely an antibacterial agent A and an antibacterial agent B, and a blank control experiment. In each set of experiments, different experimental strains were tested individually with antibacterial agents or blank controls.

In the same group, each individual experimental run was as follows: the antibacterial agents are respectively placed in the two culture dishes, the two culture dishes are placed in an inward overlapping mode, and as shown in figure 1, the non-contact antibacterial effect of the antibacterial agents is measured in a small space.

(1) Bacterial culture

For gram-positive bacteria, a cell culture tube is used for shaking culture at 40 ℃, 200 mu L of bacteria culture solution at the later stage of log phase is taken, and 1.8mL of physiological saline is added.

For other experimental strains, a single experimental strain was spread and inoculated on Potato Dextrose Agar (PDA) medium, and after one week of incubation at 30 ℃, the spores were suspended in 0.5ml of sterilized physiological saline supplemented with 0.01% SDS. The gram-positive bacteria were subjected to direct counting to determine the number of bacteria, and the other experimental strains were subjected to spore count by a hemocytometer to adjust the number of spores to about 1000/ml of suspension.

(2) Preparation of the test strains

100 μ L of the inoculation suspension was smeared in the culture medium of the experimental strain. Wherein the gram-positive bacteria use nutrient agar medium (NA); other experimental strains used potato dextrose agar medium (PDA).

(3) Preparation of antibacterial agent

For the culture of the antibacterial agent (and a blank control experiment), a nutrient agar culture medium (NA) in a culture dish is respectively covered with the antibacterial agent A and the antibacterial agent B (and is not covered with the antibacterial agent), and the culture medium inoculated with the antibacterial agent and the culture medium not inoculated (a blank control experiment) are cultured for 2 days at the temperature of 30 ℃.

(4) Culturing

And (3) mutually buckling and overlapping the experimental strain culture dish prepared in the step (2) and the antibacterial agent culture dish prepared in the step (3) and a blank control culture dish respectively, and enabling the antibacterial agent not to be in contact with the culture dish of the experimental strain, wherein the culture dish loaded with the antibacterial agent or the blank control is arranged at the lower side as shown in figure 1. The upper and lower culture dishes are fixed outside the culture dishes through medical breathable adhesive tapes. .

Culturing at 30 deg.C. The culture time is respectively as follows: the gram-positive bacteria group is 3 days, and the other experimental strains group is 7 days.

(5) Measurement of

As shown in fig. 4, the growth of the experimental strains in the experimental group was significantly inhibited compared to the control group, and it was determined that the corresponding microbial inoculum had an inhibitory effect. The following table shows the results of the antimicrobial activity tests shown in the small space. When growth inhibition was confirmed, the value was designated as + and when growth inhibition was not confirmed, the value was designated as-. The following table is recorded, and the antibacterial agents A and B have antibacterial effects on experimental strains of cladosporium cladosporioides, cladosporium globosum, alternaria, curvularia, fusarium oxysporum and rhizoctonia solani under the non-contact condition in a small space.

Table 2: results of antimicrobial activity test in a small enclosed space

Example 21.3L experiment of antibacterial Activity in closed space

In the same group, each individual experimental run was as follows: experimental strains and antibacterial agents are respectively placed in the two culture dishes and the antibacterial agents are respectively placed in the two culture dishes, the two culture dishes are placed in a 1.3L closed plastic container side by side, and as shown in figure 2, the non-contact antibacterial effect of the antibacterial agents is measured in a small space.

(1) Bacterial culture

(2) Preparation of the test strains

(3) Preparation of antibacterial agent

(4) Culturing

And (3) mutually buckling and overlapping the experimental strain culture dish prepared in the step (2) and the antibacterial agent culture dish prepared in the step (3) and the blank control culture dish respectively, and enabling the antibacterial agent not to be in contact with the culture dish of the experimental strain, wherein the culture dish loaded with the antibacterial agent or the blank control is arranged at the lower side as shown in figure 2. The upper and lower culture dishes are fixed outside the culture dishes through medical breathable adhesive tapes. .

(5) Measurement of

The calculation formula is as follows: the growth inhibition rate is represented by (average number of colonies in control group-average number of colonies in experimental strain) ÷ average number of colonies in control zone.

Compared with a control group, the growth inhibition rate of the experimental group cladosporium, cladosporium sphaeroides, alternaria alternata and rhizoctonia solani is 100%, the inhibition rate is completely inhibited, and the corresponding microbial inoculum is judged to have the inhibition effect. The following table shows the results of the antimicrobial activity tests shown in the intermediate space. The antibacterial agents A and B are proved to have antibacterial effects on the experimental strains of cladosporium cladosporioides, cladosporium sphaerothecoides, alternaria alternata, curvularia and rhizoctonia solani under the non-contact condition in a 1.3L closed space.

Table 3: test results of antibacterial activity in 1.3L of enclosed space

Example 310L test of antibacterial Activity in closed spaces

In the same group, each individual experimental run was as follows: the experimental strain and the antibacterial agent are respectively placed in the two culture dishes and the antibacterial agent is respectively placed in the two culture dishes, the two culture dishes are placed in a 10L closed plastic container side by side, and as shown in figure 3, the non-contact antibacterial effect of the antibacterial agent is measured in a small space.

(1) Bacterial culture

(2) Preparation of the test strains

(3) Preparation of antibacterial agent

(4) Culturing

And (3) mutually buckling and overlapping the experimental strain culture dish prepared in the step (2) and the antibacterial agent culture dish prepared in the step (3) and the blank control culture dish respectively, and enabling the antibacterial agent not to be in contact with the culture dish of the experimental strain, wherein the culture dish loaded with the antibacterial agent or the blank control is arranged at the lower side as shown in figure 3. The upper and lower culture dishes are fixed outside the culture dishes through medical breathable adhesive tapes. .

(5) Measurement of

Counting the average value of the diameters of the bacterial colonies in the culture dish of the experimental strain, comparing the average value of the diameters of the bacterial colonies in the control group, and calculating the bacterial colony growth inhibition rate (%), wherein the calculation formula of the bacterial colony growth inhibition rate is as follows: (1-mean value of colony diameter of treatment zone/mean value of colony diameter of control zone). 100%.

Compared with a control group, the growth of the experimental group cladosporium, cladosporium sphaerothecium, alternaria alternata and rhizoctonia solani is completely inhibited, and the corresponding microbial inoculum is judged to have the inhibiting effect. The following table shows the results of the antimicrobial activity tests shown in the large space. The antibacterial agents A and B are proved to have antibacterial effects on experimental strains of cladosporium cladosporioides, cladosporium sphaerothecum, alternaria alternata and rhizoctonia solani under the non-contact condition in a 10L closed space.

Table 4: test results of antibacterial activity in 10L enclosed space

Example 4 antibacterial Effect of antibacterial agent after Sterilization treatment

In this example, the cause and effect relationship between the growth inhibitory effect of the experimental strain and the antibacterial agent was verified, and the sterilization treatment was performed by placing the antibacterial agent in an autoclave at 120 ℃.

Two antibacterial agents A are taken for experiment, and one of the two antibacterial agents A is sterilized; and sterilizing one of the two antibacterial agents B.

The selected experimental strain is cladosporium globisporus; the other experimental procedures were the same as in example 1. The results of the experiments are shown in the table below. It can be proved that the inactivated antibacterial agent cannot play an antibacterial effect, and the growth inhibition of the experimental strain is due to the effect of the antibacterial agent.

Table 5: statistical table of antibacterial effect of antibacterial agent after sterilization treatment

Claims

1. An antibacterial method is characterized in that under the condition that an antibacterial agent is used in a closed space and is not in contact with a target object, the antibacterial agent inhibits the growth of specific bacteria; the active ingredients of the antibacterial agent are CGMCC NO: 1.270 parts by weight of spherical lysine bacillus 1-5 parts, CGMCC NO: 785 parts by weight of Bacillus thuringiensis, 250 parts by weight of yeast 150 and 600 parts by weight of photosynthetic bacteria 400; the specific bacteria are more than one of cladosporium, cladosporium sphaeroides, alternaria alternata, curvularia and rhizoctonia solani; the volume of the closed space is less than 10L; the photosynthetic bacteria are eel photosynthetic bacteria.

2. The antibacterial method according to claim 1, characterized in that: the antibacterial agent also contains poultry manure treated at a high temperature of 60-150 ℃.