CN113662007B - Preparation and application of low-temperature freezing disinfectant - Google Patents
Preparation and application of low-temperature freezing disinfectant Download PDFInfo
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Abstract
The invention provides a preparation method of a low-temperature freezing disinfectant, which comprises the following steps: s1: taking the prepared micro-nano zinc solution 500-3000ppm and placing the solution in a container; s2: adding 0.02-0.05% of double-chain quaternary ammonium salt, 10% of ethanol, 10% of glycol and 30% of anhydrous calcium chloride into the container of S1, and uniformly stirring; s3: adding purified water into the solution of S2, and uniformly stirring to obtain a low-temperature frozen disinfectant; the invention also provides application of the low-temperature freezing disinfectant, and the low-temperature freezing disinfectant is used for killing a cold-chain logistics system. The invention is non-toxic, harmless and non-irritating, has extremely high inactivation rate to most of bacteria and viruses including non-enveloped DNA virus, enveloped RNA virus, influenza A virus, pseudorabies virus and PEDV coronavirus, has the characteristics of quick killing and lasting antibiosis and antivirus, can not freeze at low temperature, and is the most applicable sterilization and killing technology of the cold-chain logistics system at present.
Description
Technical Field
The invention relates to the field of micron antibiosis, in particular to preparation and application of a low-temperature freezing disinfectant.
Background
The virus can survive for a long time in a low temperature environment (below 20 ℃ below zero), if meat and the like are frozen at 80 ℃ below zero, the virus attached to the surface of the meat can survive for at least more than half a year, and related practitioners can cause chronic infection if contacting the meat products, which leads to spreading of the virus in a latent period.
As can be seen from the above, the killing of cold chain logistics products is very urgent and is not slow. The whole cold chain logistics are sterilized and killed into four parts, namely, a person, a vehicle, food and a refrigerator, and the four parts need to be sterilized and killed respectively.
The new coronavirus is sensitive to ultraviolet rays and heat, and can effectively inactivate viruses at the temperature of 56 ℃ for 30 minutes in ester solvents such as diethyl ether, 75% ethanol, chlorine-containing disinfectants, peroxyacetic acid, chloroform and the like. Chlorhexidine is not effective in inactivating viruses. The above are traditional killing means, wherein the ultraviolet ray killing needs to be effective after lasting for 30 minutes, the heating temperature needs to be 56 ℃ for 30 minutes, and the ultraviolet ray killing and the heating temperature are not suitable for cold chain inactivation of COVID-19; ether and 75% ethanol are flammable and explosive, and are not suitable for large-area killing, particularly, ether has great harm to human bodies, and the ether is coma when inhaled by human bodies, and the ratio of LD 50: 1215mg/kg of chlorine-containing disinfectant, peracetic acid and chloroform are all corrosive and irritant substances, and can greatly harm human bodies after long-term use.
Disclosure of Invention
The invention aims to provide a preparation method and application of a low-temperature freezing disinfectant, so as to solve the problems in the background technology.
The micro-nano zinc antibacterial agent is a bacterium and virus killing spray agent detected by an authority organization of national certification, the killing rate of the micro-nano zinc antibacterial agent to various bacteria is more than 99.999 percent, the inactivation rate to non-enveloped DNA virus, enveloped RNA virus, influenza A virus, pseudorabies virus, PEDV coronavirus and the like is more than 99.99 percent, and the micro-nano zinc antibacterial agent has the characteristics of quick killing, lasting antibiosis and antivirus, and the test result shows that: LD 50: more than 5000mg/kg, which is practically nontoxic; oral inhalation LC502 h: more than 10000mg/m3 is also practical and nontoxic; the skin irritation is not stimulated for many times. Through practical application, the system can not freeze at low temperature, and is the most suitable sterilization technology for the cold-chain logistics system at present.
In order to achieve the purpose, the invention adopts the following technical scheme:
a preparation method of a low-temperature freezing disinfectant comprises the following steps:
s1: placing the prepared micro-nano zinc solution with the concentration of 500-3000ppm in a container;
s2: sequentially adding the double-chain quaternary ammonium salt, ethanol, glycol and anhydrous calcium chloride into the container of S1, and uniformly stirring;
s3: adding purified water into the solution of S2, and uniformly stirring to obtain a low-temperature frozen disinfectant;
the low-temperature freezing and disinfecting liquid comprises the following components in percentage by mass:
the balance of water;
wherein the double-chain quaternary ammonium salt is one or more of didecyl dimethyl ammonium chloride, didecyl dimethyl ammonium bromide and dioctyl dimethyl ammonium chloride.
Further, in S1, the method for preparing the micro-nano zinc solution is as follows:
step 1: preparing a zinc particle solution by using a micro-emulsion method, wherein the particle size of zinc particles is 10-600 nm;
step 2: transferring the prepared zinc particle solution to the next reaction kettle, and preparing the micro-nano zinc solution by utilizing the cavitation phenomenon.
Further, the step 1 also comprises a step 1.1: modulating the microemulsion to form a WPO (waterborne polyurethane) reverse microemulsion system; the microemulsion consists of a surfactant, a cosurfactant, an organic solvent and deionized water.
Further, the step 1.1 also comprises dissolving the surfactant in the organic solvent, mixing with the cosurfactant and deionized water, and stirring to prepare a WPO reverse microemulsion system; the surfactant is a nonionic surfactant.
Further, the volume ratio of the total volume of the surfactant, the cosurfactant and the organic solvent to the deionized water is 1-4:1, and the volume ratio of the surfactant, the cosurfactant and the organic solvent is 1-5:1: 2-4.
Further, the organic solvent is one or more of alkane and cycloalkane; the nonionic surfactant is one or more of polyoxyethylene nonyl phenyl ether, nonylphenol polyoxyethylene ether, octylphenol polyoxyethylene ether and high-carbon fatty polyoxyethylene ether; the cosurfactant is fatty alcohol; the organic solvent is cyclohexane; the cosurfactant is one or more of isoamyl alcohol, n-heptanol, n-octanol, n-nonanol, n-decanol and cetyl alcohol.
Further, the step 1 also includes a step 1.2:
respectively adding a zinc salt aqueous solution and a hydrazine hydrate solution with the concentration of 400-600 g/L into the WPO reverse microemulsion system, stirring and mixing, and reacting for 5-8h to prepare a zinc particle solution with the particle size of 10-600 nm; the reaction temperature is 40-80 ℃, and the stirring speed is 2000-5000 rpm.
Further, the volume ratio of the zinc salt aqueous solution to the hydrazine hydrate solution is 1:1, and the volume ratio of the hydrazine hydrate solution to the WPO reverse microemulsion system is 1: 3.5-4; the zinc salt is one or more of zinc sulfate, zinc nitrate, zinc citrate and zinc gluconate.
Further, the step 2 further comprises:
step 2.1: transferring the zinc particle solution prepared in the step 1 to the next reaction kettle, stirring at 60 ℃, wherein the stirring speed is 2000-5000rpm, and simultaneously introducing high-speed air flow to form a cavitation phenomenon;
step 2.2: reacting for 5 hours to prepare the micro-nano zinc solution.
The application of the low-temperature freezing disinfectant is to kill a cold-chain logistics system.
Compared with the prior art, the invention has the following advantages and beneficial effects:
the micro-emulsion method is used for preparing the micro-nano zinc particle solution, the preparation device has simple structure, the apparatus is easy to obtain, the cost is low, the preparation method is easy to operate, the zinc particles can be controlled, and the agglomeration is not easy to occur.
According to the invention, the surface of zinc particles is corroded by adopting a cavitation phenomenon to form an acute angle with an irregular shape, and when micro-nano zinc is contacted with bacteria or viruses, the micro-nano zinc shows a positive charge effect and generates coulomb force with bacterial cell walls or virus cells showing a negative charge effect; the attraction of coulomb force and the edge of the spike can kill bacteria and viruses quickly, and the bacteria and the viruses can not be propagated or transferred continuously and die; the micro-nano zinc is not consumed in the process of killing bacteria or viruses, and can continuously cause the death of the bacteria and the viruses, so the micro-nano zinc can continuously and durably kill the bacteria and the viruses, and the defect of the rapid bactericidal performance of the nano zinc is overcome.
The micro-nano zinc particle solution prepared by the invention is non-toxic, harmless, tasteless and non-irritant, can replace most disinfection products, is beneficial to environmental protection, improves the healthy living level, and achieves the effects of really, durably, effectively, stably, efficiently, environmentally and tasteless sterilization and disinfection.
The sterilization and virus killing mechanism of the micro-nano zinc solution (effective component) in the low-temperature disinfectant is as follows:
the reactivity and activity of the particle surface are enhanced due to small particle size and irregular surface shape of the micro-nano zinc, the existence range of a plurality of microorganisms is from hundreds of nanometers to tens of micrometers, the nano metal particles have larger specific surface area, higher potential energy is accumulated, the nano metal particles have nanoparticle effect, specific surface area effect and quantum tunnel effect on the whole, active oxygen ROS effect and the like, and various effects have synergistic effect to form better antibacterial activity.
When the micro-nano zinc is contacted with bacteria or viruses, the micro-nano zinc shows a positive charge effect, generates coulomb force with the bacterial cell wall or virus cell showing a negative charge effect, and can puncture the bacterial cell wall and the protein shell of the viruses to enable cytoplasm to flow out or change, so that the bacteria and the viruses cannot continue to survive or propagate;
the micro-nano zinc particles and the released zinc ions react with-NH, -COOH, -SH and the like in bacteria, so that the structural composition of the cells is damaged, the propagation of the cells is prevented, and the effect of killing the bacteria and the mold is achieved; the released zinc ions have obvious influence on the activity transfer inhibition, the amino acid metabolism and an enzyme system;
the micro-nano zinc is dissociated from dead bacteria, and a new round of sterilization is carried out repeatedly.
In addition, the micro-nano zinc induces to generate Reactive Oxygen Species (ROS), can induce oxidative stress reaction, and generates a large amount of hydroxyl radicals and hydrogen peroxide (H) 2 O 2 Leading to bacterial apoptosis.
If the edges of the micro-nano zinc similar to the spikes do not exist, only the coulomb force is not enough to puncture the cell wall in a short time, but after the surface of the micro-nano zinc is cavitated, the spikes with irregular shapes are formed, and the bacteria and the viruses can be quickly killed by the attraction of the coulomb force and the spikes, so that the bacteria and the viruses cannot be continuously propagated or transferred.
The micro-nano zinc is not consumed in the process of killing bacteria or viruses, and can continuously cause the bacteria and the viruses to die, so the micro-nano zinc can continuously and durably kill the bacteria and the viruses.
The invention is non-toxic, harmless and non-irritating, has extremely high inactivation rate to most of bacteria and viruses including non-enveloped DNA virus, enveloped RNA virus, influenza A virus, pseudorabies virus and PEDV coronavirus, has the characteristics of quick killing and lasting antibiosis and antivirus, can not freeze at low temperature, and is the most applicable sterilization and killing technology of the cold-chain logistics system at present.
Drawings
FIG. 1 is a flow chart of steps of a preparation method of a micro-nano zinc solution;
FIG. 2 is a micro-structure diagram of micro-nano zinc before cavitation effect is carried out in the invention;
FIG. 3 is a micro-structure diagram of micro-nano zinc with etching pits obtained by utilizing a cavitation effect;
FIG. 4 is a schematic diagram of the action principle of sterilization and virus killing of micro-nano zinc with corrosion pits obtained by utilizing the cavitation effect;
FIG. 5 is a particle size distribution diagram of the micro-nano zinc of the invention;
FIG. 6 is a morphological diagram of a cultured colony after the micro-nano zinc solution acts for 20s in example 2 of the invention;
FIG. 7 is a morphological diagram of a cultured colony after a micro-nano zinc solution acts for 30s in example 2 of the invention;
FIG. 8 is a morphological diagram of a cultured colony after 60s of action of a micro-nano zinc solution in example 2 of the invention
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the accompanying drawings, and it is to be understood that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and all other embodiments obtained by those skilled in the art without any inventive work based on the embodiments of the present invention belong to the scope of the present invention.
Preparation and performance examples of main component micro-nano zinc of low-temperature freezing disinfectant
Example 1
Referring to a flow chart of steps shown in fig. 1, a method for preparing micro-nano zinc capable of killing bacteria and inactivating viruses comprises the following steps:
step 1.1: dissolving 50mL of nonylphenol polyoxyethylene ether and 30mL of n-octanol in 120mL of cyclohexane, and stirring with 20mL of isoamyl alcohol and 200mL of deionized water to prepare a WPO (waterborne polyurethane emulsion) reverse microemulsion system;
step 1.2: adding 1000mL of zinc salt aqueous solution (mixed solution of zinc citrate and zinc nitrate) with the concentration of 450g/L and 1000mL of hydrazine hydrate solution into 3600mL of WPO reverse microemulsion for mixing, and reacting for 6.5h at the rotation speed of 65℃ and 4600rpm to prepare zinc particle solution with the particle size of 10-600 nm;
step 2.1: transferring the prepared zinc particle solution into the next reaction kettle, stirring at 60 ℃, 4600rpm, and introducing high-speed air flow to form a cavitation phenomenon;
step 2.2: the reaction time is 5 hours, and the micro-nano zinc solution is prepared.
A TEM image of the prepared zinc particles is shown in fig. 2, and a TEM image of the prepared micro-nano zinc is shown in fig. 3.
As shown in fig. 5, the microemulsion method is used to prepare zinc with particle size between micron and nanometer, and the reaction conditions are controlled to prepare zinc particles with particle size:
preferred results are:
the zinc particles (2) account for 50% of the total, and the particle diameter of the zinc particles
Accounting for 30% of the total.
Example 2
The micro-nano zinc solution prepared in the example 1 is prepared into a micro-nano zinc aqueous solution with the concentration of 1000mg/kg, and then the micro-nano zinc aqueous solution is sequentially diluted into a solution with the concentration of: the method comprises the following steps of sequentially marking micro-nano zinc aqueous solutions with the concentrations of 800mg/kg, 500mg/kg, 400mg/kg and 300mg/kg as a group 5, a group 4, a group 3, a group 2 and a group 1, measuring the sterilization rate of the micro-nano zinc aqueous solution according to a suspension quantitative method of disinfection technical specification, wherein test strains comprise escherichia coli, staphylococcus aureus, candida albicans and pseudomonas aeruginosa for 20s, 30s and 60 s; the test results are shown in table 1, the morphologies of the cultured colonies after the micro-nano zinc aqueous solutions with different concentrations in the groups 1 to 5 act on escherichia coli, staphylococcus aureus, candida albicans and pseudomonas aeruginosa for 20s are shown in fig. 6, the morphologies of the cultured colonies after the micro-nano zinc aqueous solutions act on escherichia coli, staphylococcus aureus, candida albicans and pseudomonas aeruginosa for 30s are shown in fig. 7, and the morphologies of the cultured colonies after the micro-nano zinc aqueous solutions act on escherichia coli, staphylococcus aureus, candida albicans and pseudomonas aeruginosa for 60s are shown in fig. 8.
TABLE 1 Sterilization effect of micro-nano zinc solution under different time
Example 3
The micro-nano zinc solution prepared in the example 1 is prepared into a micro-nano zinc aqueous solution with the concentration of 1000mg/kg, and then the micro-nano zinc aqueous solution is sequentially diluted into a solution with the concentration of: the method comprises the following steps of sequentially marking micro-nano zinc aqueous solutions with the concentrations of 1000mg/kg, 800mg/kg, 500mg/kg, 400mg/kg and 300mg/kg as a group 5, a group 4, a group 3, a group 2 and a group 1, determining the virus inactivation rate of the micro-nano zinc aqueous solution according to disinfection technical specifications, testing the viruses to be poliovirus, influenza A virus H1N1, enterovirus and avian influenza virus H5N1 for 10s, 20s and 30 s; the test results are shown in table 2.
TABLE 2 Virus inactivating effect of micro-nano zinc solution at different time
Diluting 3% hydrogen peroxide disinfectant sold in the market into the following concentrations in sequence: the method comprises the steps of (1) sequentially marking 1000mg/kg, 800mg/kg, 500mg/kg, 400mg/kg and 300mg/kg of hydrogen peroxide aqueous solution with the concentrations of 1000mg/kg, 800mg/kg, 500mg/kg, 400mg/kg and 300mg/kg of hydrogen peroxide aqueous solution as a group 5, a group 4, a group 3, a group 2 and a group 1, determining the virus inactivation rate of the hydrogen peroxide aqueous solution according to the disinfection technical specification, and testing the viruses to be poliovirus, influenza A virus H1N1, enterovirus and avian influenza virus H5N1 for 10s, 20s and 30s respectively; the test results are shown in table 3.
TABLE 3 Effect of hydrogen peroxide on inactivation of virus at different times
Example 4
Preparing the prepared micro-nano zinc into an aqueous solution with the concentration of 1000mg/kg, diluting the micro-nano zinc aqueous solution by 10000 times to obtain the micro-nano zinc aqueous solution with the concentration of 100 mu g/kg, and taking 5 parallel samples to mark as: group 1, group 2, group 3, group 4 and group 5, groups 1 to 5 respectively with a concentration of 10 6.0 TCID 50 Mixing pseudorabies virus and coronavirus (PEDV) in a contact manner for 30min, and measuring the virus inactivation rate; the test results are shown in table 4.
TABLE 4 Virus inactivating effect of micro-nano zinc solution
Diluting commercially available glutaraldehyde disinfection solution to obtain glutaraldehyde aqueous solution with the concentration of 100 mug/kg, and taking 5 parallel samples as markers: group 1, group 2, group 3, group 4 and group 5, groups 1 to 5 respectively with a concentration of 10 6.0 TCID 50 Mixing pseudorabies virus and coronavirus (PEDV) in a contact manner for 30min, and measuring the virus inactivation rate; the test results are shown in table 5.
TABLE 5 Effect of glutaraldehyde inactivation of viruses
According to the experimental data of the embodiment, the micro-nano zinc solution prepared by the method can realize the effect of quick sterilization.
Referring to fig. 4, the micro-nano zinc solution prepared by the invention has a sterilization and virucidal mechanism:
the reactivity and activity of the particle surface are enhanced due to small particle size and irregular surface shape of the micro-nano zinc, the existence range of a plurality of microorganisms is from hundreds of nanometers to tens of micrometers, the nano metal particles have larger specific surface area, higher potential energy is accumulated, the nano metal particles have nanoparticle effect, specific surface area effect and quantum tunnel effect on the whole, active oxygen ROS effect and the like, and various effects have synergistic effect to form better antibacterial activity.
When the micro-nano zinc is contacted with bacteria or viruses, the micro-nano zinc shows a positive charge effect, generates coulomb force with the bacterial cell wall or virus cell showing a negative charge effect, and can puncture the bacterial cell wall and the protein shell of the viruses to enable cytoplasm to flow out or change, so that the bacteria and the viruses cannot continue to survive or propagate;
the micro-nano zinc particles and the released zinc ions react with-NH, -COOH, -SH and the like in bacteria, so that the structural composition of the cells is damaged, the propagation of the cells is prevented, and the effect of killing the bacteria and the mold is achieved; the released zinc ions have obvious influence on the activity transfer inhibition, the amino acid metabolism and an enzyme system;
the micro-nano zinc is dissociated from dead bacteria, and a new round of sterilization is carried out repeatedly.
In addition, the micro-nano zinc induces to generate Reactive Oxygen Species (ROS), can induce oxidative stress reaction, and generates a large amount of hydroxyl radicals and hydrogen peroxide (H) 2 O 2 Leading to bacterial apoptosis.
If the edges of the micro-nano zinc similar to the spikes do not exist, only the coulomb force is not enough to puncture the cell wall in a short time, but after the surface of the micro-nano zinc is cavitated, the spikes with irregular shapes are formed, and the bacteria and the viruses can be quickly killed by the attraction of the coulomb force and the spikes, so that the bacteria and the viruses cannot be continuously propagated or transferred.
The micro-nano zinc is not consumed in the process of killing bacteria or viruses, and can continuously cause the bacteria and the viruses to die, so the micro-nano zinc can continuously and durably kill the bacteria and the viruses.
Preparation example, toxicity test example and application example of low-temperature freezing disinfectant
Example 5
A preparation method of a low-temperature freezing disinfectant comprises the following steps:
s1: placing the micro-nano zinc solution prepared in the embodiment 1 in a container;
s2: sequentially adding the double-chain quaternary ammonium salt, ethanol, glycol and anhydrous calcium chloride into the container of S1, and uniformly stirring;
s3: adding purified water into the solution of S2, and uniformly stirring to obtain a low-temperature frozen disinfectant;
the low-temperature freezing and disinfecting liquid comprises the following components in percentage by mass:
the application of low-temperature freezing disinfectant is used for killing a cold-chain logistics system.
Examples 6 to 9 are safety tests of low-temperature freezing disinfectant
Example 6
Acute oral toxicity test (mouse)
1. Materials and animals
And (3) testing a sample: 5000mg of the low-temperature frozen disinfectant prepared in example 5 was weighed, and purified water was added to 20ml to prepare a test substance.
Animal and feed SPF grade healthy ICR mice 20, half male and half female, weight 18.2-21.8 g.
The test conditions are as follows: ambient temperature: 22 + -2 deg.C, relative humidity 40-70%, main apparatus: electronic scale, electronic balance
2. Method of producing a composite material
The detection basis is as follows: refer to the Disinfection Specification, 2002 edition, second section, Disinfection product testing Specification, 2.3.1 acute oral toxicity test, Ministry of health.
The experimental method comprises the following steps: one maximum test. The designed dose is 5000mg/kg b.wt., after the mice are fasted overnight, the mice are taken through oral one-time gavage, the gavage volume is 20ml/kg b.wt, and the mice are taken 4h after gavage, and the observation period is 14 d.
3. And (3) test results:
after the gavage of the male and female mice, no obvious poisoning phenomenon is seen, no death is caused in the observation period, no obvious abnormality is seen in the gross anatomy after the experiment is finished, and the death result of the experimental animals is shown in a table 6:
TABLE 6 mouse acute oral toxicity test results
And (4) conclusion: the acute oral LD50 value of the sample to female and male mice is more than 5000mg/kg b.wt., which belongs to the actual nontoxic grade.
Example 7
Multiple complete skin irritation test
1. Materials and animals
And (3) testing a sample: the stock solution of the cryo-disinfectant prepared in example 5 was used as a test substance.
Animals: 4 healthy new Zealand white rabbits without skin disease, the weight of the ordinary rabbit is 2.4-2.9kg
The test conditions are as follows: the ambient temperature is 22 plus or minus 2 ℃, and the relative humidity is 40-70%; the main apparatus is as follows: baby scale
2. Method for producing a composite material
The inspection basis is as follows: the second part of the "Disinfection Specification" of the Ministry of health (2002) 2.3.3 skin irritation tests.
The test method comprises the following steps: on the day before the test, the hair on both sides of the dorsal spine of the rabbit was cut off at a range of 3cmx3 cm. During test, about 0.5ml of the test substance is coated on the unhaired skin on one side, the skin on the other side is used as a blank control, the coating area is 2.5cmx2.5cm, the test substance is coated once a day and is continuously coated for 14 d. After 4h of application, the paint was rinsed with warm water to remove residues, and after 24h the results were observed, scored according to Table 2-11 in Disinfection Specification (2002 edition), and the control area was treated as in the experimental area. At the end of the experiment, skin irritation intensity was graded as per Table 2-12 in Disinfection Specification (2002 edition).
3. Test results
See Table 7 below, without irritation and other toxic effects
TABLE 7 test results of repeated intact skin irritation of test substances to rabbits
And (4) conclusion: the integral average value of the multiple complete skin irritation reaction of the sample to the rabbit is 0, and the irritation strength is nonirritant.
Example 8
Skin allergy test
1. Materials and animals
And (3) testing a sample: the stock solution of the cryo-disinfectant prepared in example 5 was used as a test substance.
Animals: 32 healthy white guinea pigs with a weight of 200-.
The test conditions are as follows: ambient temperature 22 ± 2 ℃, relative humidity: 40 to 70 percent.
2. The method comprises the following steps:
the inspection basis is as follows: the second part 2.3.6 of the Ministry of health (2002 edition) Disinfection Specification is the skin allergy test.
The test method comprises the following steps: local seal coating (BT) method. Animals were randomly divided into experimental groups and negative control groups of 16 animals each. The back of the skin is dehaired and preserved by 3cmx3cm 24h before each coat. Sensitization induction: applying 0.5ml of the test substance on the left skin-removing area of the experimental group, covering with 2 layers of gauze, sealing and fixing with adhesive plaster for 6h, and washing off the test substance. Skin reactions were observed after 24h and 48 h.
Positive control: 2, 4-dinitrochlorobenzene was used as a positive control, the induction and excitation concentrations were 2.0% (w/v) and 1.0% (w/v), respectively, and the test procedure was the same as that of the test group.
3. Test results
No skin reactions were observed with topical occlusive dressings, as shown in Table 8.
TABLE 8 results of skin allergy (sensitization) test in guinea pigs
Note: the skin erythema reaction intensity and edema reaction intensity are animal ratio proportion examples under the condition that the skin reaction integral is 0, 1, 2, 3 and 4
And (4) conclusion: the skin sensitization rate (%) of the sample to the test animal guinea pig was 0, and no skin allergy was observed.
Example 9
Low temperature test
The inspection basis is as follows:
the test is carried out according to the technical requirement for evaluating the sanitation and safety of the low-temperature disinfectant (State and health supervision letters [ 2020 ] 1062).
Evaluation basis:
the evaluation is carried out according to the technical requirement for evaluating the sanitation and safety of the low-temperature disinfectant (State and health supervision letters [ 2020 ] 1062).
And (4) checking and concluding:
the low-temperature test result meets the qualified regulation of the technical requirement for evaluating the sanitation and safety of the low-temperature disinfectant (No. 1062 of State health supervision letters (2020)).
1. Equipment:
a low temperature disinfectant.
Refrigerator (No. LH 078).
Pipette (5mL), centrifuge tube 10 mL).
2. The method comprises the following steps:
the detection basis is as follows: the technical requirement for evaluating the sanitary safety of the low-temperature disinfectant (State health supervision letters [ 2020 ] 1062).
The test method comprises the following steps: 5mL of low-temperature disinfectant is placed at the constant temperature of-18 ℃ overnight (>8h), and the properties of the disinfectant are observed and recorded.
3. As a result:
the low temperature disinfectant is stored at-18 ℃ overnight (>8h), and the sample keeps liquid state without precipitation and crystallization.
And (4) conclusion: through detection, the low-temperature test result of the low-temperature disinfectant meets the qualified regulation of the technical requirement for evaluating the sanitation and safety of the low-temperature disinfectant (State administration and supervision letters [ 2020 ] 1062).
Example 10
And (3) low-temperature freezing disinfection solution sterilization test:
basis of examination
The test was carried out according to the Disinfection Specification (2002 edition) items 2.2.1.2.14, 2.2.1.4, 2.1.1.5.6, 2.1.1.7.5(1) and 21.2.9.
Basis of evaluation
The evaluation is carried out according to the technical Specification for disinfection (2002 edition) and the technical requirement for evaluating the sanitation and safety of the low-temperature disinfectant (State health supervision letters [ 2020 ] 062).
Inspection conclusion microbial indicator of killing
1. After 3 times of repeated tests, under the constant temperature condition of-18 ℃, the neutralizing agent solution of the D/E neutralizing broth can effectively neutralize the residual action of the low-temperature disinfectant stored at-18 ℃ overnight (more than 8h) on staphylococcus aureus, and the neutralizing agent and a neutralizing product thereof have no adverse effect on a culture medium and basically have no effect on the growth of the staphylococcus aureus.
2. After 3 repeated tests, the low-temperature disinfectant stored at-18 ℃ overnight (more than 8 hours) is applied under the constant temperature condition of-18 ℃ for 30.0min, the killing log values of staphylococcus aureus on the cloth carrier are all more than 3.00, and the requirements of disinfection qualification in disinfection technical Specification (2002 edition) and microorganism killing tests in Low-temperature disinfectant sanitation evaluation technical requirement (national defense supervision letter [ 2020 ] 1062) are met.
3. After 3 times of repeated tests, the low-temperature disinfectant stored at-18 ℃ overnight (more than 8h) is applied at the constant temperature of-18 ℃ for 30.0min, and the killing log values of the Escherichia coli on the cloth carrier are all more than 3.00. Meets the requirements of disinfection technical specification (2002 edition) disinfection qualification and low-temperature disinfectant sanitation safety evaluation technical requirement (national defense supervision letter [ 2020 ] 1062) microorganism killing test
4. After 3 repeated tests, the low-temperature disinfectant stored at-18 ℃ overnight (more than 8h) is applied to the refrigerated storage environment condition at the temperature of-18.0 ℃ for spray disinfection, the action is 30.0min, the killing logarithm value of staphylococcus aureus on the cloth carrier at the position which is most difficult to disinfect is more than 3.00, and the sterilization specification accords with the disinfection specification (2002 edition) disinfection qualification and the technical requirement for evaluating the sanitation safety of the low-temperature disinfectant (national defense supervision letter [ 2020 ] 1062) on-site test
5. After 3 repeated tests, the low-temperature disinfectant stored at-18 ℃ overnight (more than 8 hours) is applied to the cold storage environment condition at the temperature of-18.0 ℃ for spray disinfection, the action is 30.0min, the killing logarithm value of escherichia coli on the cloth carrier at the position which is most difficult to disinfect is more than 3.00, and the sterilization specification accords with the sterilization specification (2002 edition) disinfection qualification and the technical requirement for the low-temperature disinfectant sanitation safety evaluation (national defense supervision letter [ 2020 ] 1062) field test specification.
Quantitative killing test for staphylococcus aureus carrier
1. Equipment:
test strains: staphylococcus aureus (ATCC 6538), passage 6;
a low temperature disinfectant;
neutralizing agent: D/E neutralization broth;
diluting liquid: tryptone saline solution (TPS);
tryptone soy agar medium (TSA), tryptone soy broth medium (TSB);
test carrier: sterile square plain cotton sheets (10 mm. times.10 mm);
biological safety cabinet, incubator, thermostat, refrigerator. Sterile equipment, electronic timers, and the like.
2. The method comprises the following steps:
detecting basis; "Disinfection Specification" 2002 edition) items 2.1.1.5.6 and 2.1.1.7.5 (1).
Preparing a bacterial carrier: preparing bacterial suspension for bacterial infection by using TSB, sucking 10 mu L of bacterial suspension by using a micropipette, dripping and dyeing a dry carrier, uniformly coating, and drying in an incubator at 37 ℃ for 30min to prepare a bacterial carrier (hereinafter referred to as bacterial tablet). Standing the dried mycelia at-18 deg.C for 30 min.
And (3) identification test of a neutralizer: the low-temperature disinfectant stored at-18 ℃ overnight (>8h) is used for the test, the action time is 2.5min under the constant temperature condition of-18 ℃, and the test is repeated for 3 times.
And (3) sterilization test: the experiment was repeated 3 times with low temperature disinfectant stored overnight (more than 8h) at-18 deg.C for 15.0min, 30.0min and 45.0min at constant temperature of-18 deg.C.
3. As a result: killing effect on staphylococcus aureus
After 3 times of repeated tests, the low-temperature disinfectant stored at-18 ℃ overnight (> 3h) is applied under the constant temperature condition of-18 ℃ to act for 30.0nin, and the killing log values of the staphylococcus aureus on the cloth carrier are all more than 3.00, which is shown in Table 9.
TABLE 9 killing effect on Staphylococcus aureus
Second, quantitative killing test of colibacillus carrier
1. Equipment:
test strains: escherichia coli (8099), passage 6;
a low temperature disinfectant;
neutralizing agent: D/E neutralization broth;
diluting liquid: tryptone saline solution (TPS);
tryptone soy agar medium (TSA), tryptone soy broth medium (TSB);
test carrier: sterile square float tabby (10 mm. times.10 mm);
biosafety cabinets, incubators, thermostats, refrigerators, sterile equipment, electronic timers, and the like.
2. The method comprises the following steps:
and (6) detection basis. The specification for disinfection (2002 edition) section 2.1.1.7.5.
Preparing a bacterial carrier: TSB is used for preparing bacterial suspension for bacteria infection, a micropipette sucks 10 mu L of bacterial suspension, and the bacterial suspension is dripped into a carrier. Uniformly spreading, and drying at 37 deg.C for 30min to obtain bacterial carrier (hereinafter referred to as bacterial tablet). Standing the dried mycelia at-18 deg.C for 30 min.
And (3) sterilization test: the experiment was repeated 3 times with a low temperature disinfectant stored overnight (>8h) at-18 ℃ for 15.0xin, 30.0min and 45.0min at a constant temperature of-18 ℃.
3. As a result: the killing effect on escherichia coli;
after 3 repeated experiments, the low-temperature disinfectant stored at-18 ℃ overnight (more than 8h) is applied at the constant temperature of-18 ℃ for 30.0min, and the killing log values of escherichia coli on the cloth carrier are all more than 3.00, which is shown in Table 10.
TABLE 10 effect on Escherichia coli
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, and that the preferred embodiments of the invention and described in the specification are only preferred embodiments of the invention and are not intended to limit the invention, and that various changes and modifications may be made without departing from the novel spirit and scope of the invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (7)
1. The preparation method of the low-temperature freezing disinfectant is characterized by comprising the following steps:
s1: placing the prepared micro-nano zinc solution with the concentration of 500-3000ppm in a container; the preparation method of the micro-nano zinc solution comprises the following steps: step 1: preparing a WPO (WPO) reverse microemulsion system from a microemulsion, wherein the microemulsion is composed of a nonionic surfactant, a cosurfactant, an organic solvent and deionized water;
step 2: respectively adding a zinc salt aqueous solution and a hydrazine hydrate solution with the concentration of 400-600 g/L into the WPO reversed-phase microemulsion system obtained in the step (1), stirring and mixing, and reacting for 5-8h to obtain a zinc particle solution with the particle size of 10-600 nm; the reaction temperature is 40-80 ℃, and the stirring speed is 2000-5000 rpm;
and step 3: transferring the zinc particle solution prepared in the step 2 to the next reaction kettle, stirring at 60 ℃, wherein the stirring speed is 2000-plus 5000rpm, introducing high-speed air flow, and preparing the micro-nano zinc solution by utilizing the cavitation phenomenon;
s2: sequentially adding the double-chain quaternary ammonium salt, ethanol, glycol and anhydrous calcium chloride into the container of S1, and uniformly stirring;
s3: adding purified water into the solution of S2, and uniformly stirring to obtain a low-temperature frozen disinfectant;
the low-temperature freezing and disinfecting liquid comprises the following components in percentage by mass:
0.5 percent of micro-nano zinc solution
0.2 to 0.5 percent of double-chain quaternary ammonium salt
Ethanol content is 10%
10 percent of ethylene glycol
30 percent of anhydrous calcium chloride
The balance of water;
wherein the double-chain quaternary ammonium salt is one or more of didecyl dimethyl ammonium chloride, didecyl dimethyl ammonium bromide and dioctyl dimethyl ammonium chloride.
2. The method for preparing a low-temperature freeze disinfectant as claimed in claim 1, wherein the step 1 further comprises dissolving the non-ionic surfactant in the organic solvent, mixing with the co-surfactant and deionized water, and stirring to prepare the WPO reverse microemulsion system.
3. The method for preparing a low-temperature freeze disinfectant as claimed in claim 2, wherein the volume ratio of the total volume of the nonionic surfactant, the cosurfactant and the organic solvent to the deionized water is 1-4:1, and the volume ratio of the nonionic surfactant, the cosurfactant and the organic solvent is 1-5:1: 2-4.
4. The method for preparing a low-temperature freeze-thaw liquid according to claim 3, wherein the organic solvent is one or more of alkane and cycloalkane; the nonionic surfactant is one or more of polyoxyethylene nonyl phenyl ether, nonylphenol polyoxyethylene ether, octylphenol polyoxyethylene ether and high-carbon fatty polyoxyethylene ether; the cosurfactant is fatty alcohol; the organic solvent is cyclohexane; the cosurfactant is one or more of isoamyl alcohol, n-heptanol, n-octanol, n-nonanol, n-decanol and cetyl alcohol.
5. The preparation method of a low-temperature freezing disinfectant according to claim 4, wherein the volume ratio of the zinc salt aqueous solution to the hydrazine hydrate solution is 1:1, and the volume ratio of the hydrazine hydrate solution to the WPO reverse microemulsion system is 1: 3.5-4; the zinc salt is one or more of zinc sulfate, zinc nitrate, zinc citrate and zinc gluconate.
6. The method as claimed in claim 1, wherein the step 3 further comprises:
step 3.1: transferring the zinc particle solution prepared in the step 2 to the next reaction kettle, stirring at 60 ℃, wherein the stirring speed is 2000-5000rpm, and simultaneously introducing high-speed air flow to form a cavitation phenomenon;
step 3.2: reacting for 5 hours to prepare the micro-nano zinc solution.
7. Use of a cryo-disinfectant prepared by the process for the preparation of a cryo-disinfectant according to claims 1-6, wherein the cryo-disinfectant is used for the disinfection of cold chain logistics systems.
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| JP3817886B2 (en) * | 1998-02-12 | 2006-09-06 | 大日本インキ化学工業株式会社 | Plate making method of direct drawing type lithographic printing original plate |
| US6878309B2 (en) * | 2001-02-12 | 2005-04-12 | Dober Chemical Corp. | Controlled release cooling additive composition |
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| WO2006062846A2 (en) * | 2004-12-09 | 2006-06-15 | The Dial Corporation | Compositions having a high antiviral and antibacterial efficacy |
| CN101475206A (en) * | 2009-01-13 | 2009-07-08 | 东华大学 | Method for preparing ZnO nanorod with controllable distribution by growing in microchannel |
| CN110179681A (en) * | 2019-04-26 | 2019-08-30 | 得洁日化科技有限公司 | A kind of nano silver shower cream and preparation method thereof for capableing of antibiotic and sterilizing |
| CN111657299A (en) * | 2020-07-10 | 2020-09-15 | 广州佳乐生态科技有限公司 | Composite nano-ion bacteriostatic agent |
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