CN115918657A - Preparation method of glutaraldehyde decamethylammonium bromide solution - Google Patents
Preparation method of glutaraldehyde decamethylammonium bromide solution Download PDFInfo
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- CN115918657A CN115918657A CN202211540720.8A CN202211540720A CN115918657A CN 115918657 A CN115918657 A CN 115918657A CN 202211540720 A CN202211540720 A CN 202211540720A CN 115918657 A CN115918657 A CN 115918657A
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- glutaraldehyde
- bromide solution
- decamethylammonium bromide
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- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 title claims abstract description 107
- CPELXLSAUQHCOX-UHFFFAOYSA-M Bromide Chemical compound [Br-] CPELXLSAUQHCOX-UHFFFAOYSA-M 0.000 title claims abstract description 83
- 238000002360 preparation method Methods 0.000 title claims abstract description 19
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims abstract description 59
- 239000000203 mixture Substances 0.000 claims abstract description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 239000008367 deionised water Substances 0.000 claims abstract description 15
- 229910021641 deionized water Inorganic materials 0.000 claims abstract description 15
- 229940051841 polyoxyethylene ether Drugs 0.000 claims abstract description 15
- 229920000056 polyoxyethylene ether Polymers 0.000 claims abstract description 15
- 238000002156 mixing Methods 0.000 claims abstract description 13
- 239000002994 raw material Substances 0.000 claims abstract description 10
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 7
- 238000003756 stirring Methods 0.000 claims abstract description 7
- 238000005303 weighing Methods 0.000 claims abstract description 7
- 239000000243 solution Substances 0.000 claims description 88
- 238000010790 dilution Methods 0.000 claims description 32
- 239000012895 dilution Substances 0.000 claims description 32
- 241000701093 Suid alphaherpesvirus 1 Species 0.000 claims description 18
- 241001135989 Porcine reproductive and respiratory syndrome virus Species 0.000 claims description 17
- SWLVFNYSXGMGBS-UHFFFAOYSA-N ammonium bromide Chemical compound [NH4+].[Br-] SWLVFNYSXGMGBS-UHFFFAOYSA-N 0.000 claims description 17
- 241000700605 Viruses Species 0.000 claims description 16
- 239000007788 liquid Substances 0.000 claims description 14
- 238000004659 sterilization and disinfection Methods 0.000 claims description 14
- 238000000034 method Methods 0.000 claims description 13
- 238000012423 maintenance Methods 0.000 claims description 12
- 241000222122 Candida albicans Species 0.000 claims description 11
- 241000588724 Escherichia coli Species 0.000 claims description 11
- 241000191967 Staphylococcus aureus Species 0.000 claims description 11
- 229940095731 candida albicans Drugs 0.000 claims description 11
- 230000001717 pathogenic effect Effects 0.000 claims description 10
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- 235000014469 Bacillus subtilis Nutrition 0.000 claims description 8
- 238000007865 diluting Methods 0.000 claims description 5
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- 238000002474 experimental method Methods 0.000 claims description 4
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- 239000013642 negative control Substances 0.000 claims description 3
- 230000003203 everyday effect Effects 0.000 claims description 2
- 238000001179 sorption measurement Methods 0.000 claims 1
- 239000000645 desinfectant Substances 0.000 abstract description 15
- 230000001954 sterilising effect Effects 0.000 abstract description 5
- 238000003860 storage Methods 0.000 abstract description 4
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 26
- 238000012360 testing method Methods 0.000 description 26
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 20
- 235000011187 glycerol Nutrition 0.000 description 16
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 11
- 230000000694 effects Effects 0.000 description 10
- 230000000052 comparative effect Effects 0.000 description 8
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 6
- -1 quaternary ammonium salt cations Chemical class 0.000 description 6
- 150000002191 fatty alcohols Chemical class 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 229910000831 Steel Inorganic materials 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 239000000463 material Substances 0.000 description 4
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- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 3
- 150000001299 aldehydes Chemical class 0.000 description 3
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- 102000004169 proteins and genes Human genes 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 150000003242 quaternary ammonium salts Chemical class 0.000 description 3
- 238000013112 stability test Methods 0.000 description 3
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- 241000282693 Cercopithecidae Species 0.000 description 2
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- 230000008014 freezing Effects 0.000 description 2
- 238000007710 freezing Methods 0.000 description 2
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- 229910052751 metal Inorganic materials 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 239000002736 nonionic surfactant Substances 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 241000699800 Cricetinae Species 0.000 description 1
- 241001473385 H5N1 subtype Species 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 125000003172 aldehyde group Chemical group 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 230000002528 anti-freeze Effects 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- GDTBXPJZTBHREO-UHFFFAOYSA-N bromine Substances BrBr GDTBXPJZTBHREO-UHFFFAOYSA-N 0.000 description 1
- 229910052794 bromium Inorganic materials 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
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- 230000012447 hatching Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000007794 irritation Effects 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
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- 125000002467 phosphate group Chemical group [H]OP(=O)(O[H])O[*] 0.000 description 1
- 150000003904 phospholipids Chemical class 0.000 description 1
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- 231100000331 toxic Toxicity 0.000 description 1
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N35/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having two bonds to hetero atoms with at the most one bond to halogen, e.g. aldehyde radical
- A01N35/02—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having two bonds to hetero atoms with at the most one bond to halogen, e.g. aldehyde radical containing aliphatically bound aldehyde or keto groups, or thio analogues thereof; Derivatives thereof, e.g. acetals
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/02—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests containing liquids as carriers, diluents or solvents
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/22—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests containing ingredients stabilising the active ingredients
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/30—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests characterised by the surfactants
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N33/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic nitrogen compounds
- A01N33/02—Amines; Quaternary ammonium compounds
- A01N33/12—Quaternary ammonium compounds
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The invention provides a preparation method of a glutaraldehyde decamethylammonium bromide solution, belonging to the technical field of disinfectants, wherein the glutaraldehyde decamethylammonium bromide solution is prepared from the following raw material components in percentage by weight: 4-7% of glutaraldehyde, 4-7% of decamethylammonium bromide, 0.2-2% of high-carbon fatty alcohol polyoxyethylene ether, 6-20% of glycerol and the balance of deionized water. The preparation method comprises the following steps: s1, weighing; s2, mixing glutaraldehyde and ammonium decamethyl bromide to obtain a mixture A; s3, adding the fatty alcohol-polyoxyethylene ether and the glycerol into deionized water, then adding the mixture A, and stirring and uniformly mixing to obtain a mixture B; and S4, adjusting the pH value of the mixture B obtained in the step S2, and adding deionized water to a constant volume to obtain a glutaraldehyde decamethylammonium bromide solution. The glutaraldehyde decamethylammonium bromide solution can be applied to the field of disinfectants. The invention has no pungent smell, high stability, long storage period and obvious sterilizing effect at low temperature.
Description
The application is a divisional application of a patent application named as 'a glutaraldehyde decamethylammonium bromide solution and a preparation method thereof', the application date of the original application is 12 months and 10 days in 2020, and the application number is 202011432269.9.
Technical Field
The invention relates to the technical field of disinfectants, in particular to a preparation method of a glutaraldehyde decamethylammonium bromide solution.
Background
Glutaraldehyde is a high-efficiency disinfectant, and free aldehyde groups of glutaraldehyde bind to amino groups of proteins or enzymes on or in the cell surface to cause a series of reactions, resulting in the death of microorganisms. The reaction rate of the glutaraldehyde, the protein and the enzyme depends on the pH value of the solution, the reaction rate is increased along with the increase of the pH value within the range of pH4-9, the sterilization effect of the alkaline glutaraldehyde is obviously stronger than that of the acidic glutaraldehyde, and particularly, the difference is larger when the temperature is lower. However, alkaline glutaraldehyde is unstable, easily polymerizes into polymers, and easily loses activity.
The decamethyl ammonium bromide is double-chain quaternary ammonium salt, belongs to an outer membrane active compound, can dissociate out quaternary ammonium salt cations in a solution state, is combined with phosphate groups with negative charges in bacterial cytoplasmic membrane phospholipid, has a bacteriostatic action at low concentration and has a bactericidal action at high concentration. The bromine ions greatly increase the hydrophilicity and lipophilicity of molecules, can rapidly permeate into a plasma membrane lipid layer and a protein layer, change the permeability of the membrane and achieve the bactericidal effect.
The glutaraldehyde decamethyl ammonium bromide solution is a novel compound disinfectant newly introduced in recent years, integrates the advantages of aldehyde disinfectant and quaternary ammonium salt, has a double-sterilization effect due to the mutual synergy of the aldehyde disinfectant and the quaternary ammonium salt, is a broad-spectrum, high-efficiency and low-toxicity aldehyde disinfectant, has no corrosivity on various materials such as metal, glass, rubber, plastic and the like, has small irritation on skin and mucosa, and has strong penetrability and stable performance. The common glutaraldehyde decamethylammonium bromide exists in the form of hydrate (mostly cyclic structure hydrate), so that glutaraldehyde is volatilized in a short time due to the evaporation of water, and the effective killing time is greatly shortened. The Chinese patent application CN201810637946.7 discloses a glutaraldehyde decamethylammonium bromide solution and a preparation method thereof, and the disinfectant mainly comprises glutaraldehyde and decamethylammonium bromide, has the characteristics of no pungent smell, high stability and long-term storage, but does not consider the use under low temperature conditions for treatment. Chinese patent application CN201911421388.1 discloses a glutaraldehyde decamethylammonium bromide compound disinfectant suitable for low-temperature use, which consists of glutaraldehyde, decamethylammonium bromide, a deicing agent and a solvent, wherein the deicing agent adopts ethanol, ethylene glycol and sodium chloride. However, ethanol and ethylene glycol are volatile and ethylene glycol is toxic. And the problem of metal corrosion cannot be thoroughly solved when ethanol, glycol or sodium chloride is used.
In northern areas, the time is long in winter, the climate is cold, the disinfection effect of the disinfectant can be influenced in a low-temperature environment, and the freezing of the disinfection solution can greatly reduce the disinfection result of the disinfectant and even make the disinfectant ineffective. Therefore, there is an urgent need for a glutaraldehyde decamethylammonium bromide solution that has reduced pungent odor, high stability, and can maintain a significant sterilizing effect even in a low-temperature environment.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide a preparation method of a glutaraldehyde decamethylammonium bromide solution.
In order to achieve the purpose, the invention provides the following scheme:
a preparation method of a glutaraldehyde decamethylammonium bromide solution comprises the following steps:
s1, weighing: weighing raw materials, wherein the raw materials comprise 4-7% of glutaraldehyde, 4-7% of decamethylammonium bromide, 0.2-2% of high-carbon fatty alcohol polyoxyethylene ether, 6-20% of glycerol and the balance of deionized water;
s2, preparation of a mixture A: mixing glutaraldehyde and decamethylammonium bromide, and stirring uniformly to obtain a mixture A;
s3, preparation of a mixture B: adding fatty alcohol-polyoxyethylene ether and glycerol into 60ml of deionized water, then adding the mixture A, and stirring and uniformly mixing to obtain a mixture B;
s4, preparing a glutaraldehyde decamethyl ammonium bromide solution: adjusting the pH value of the mixture B obtained in the step S2 to 5.4-6.6, adding deionized water and fixing the volume to 100ml to obtain a glutaraldehyde decamethylammonium bromide solution;
the glutaraldehyde decamethyl ammonium bromide solution is used for carrying out dilution disinfection with different concentrations according to use scenes;
the glutaraldehyde decamethylammonium bromide solution has the maximum dilution multiple of 1 for 100% killing of escherichia coli, staphylococcus aureus and candida albicans, and has the maximum dilution multiple of 1 for 100% killing of bacillus subtilis.
Preferably, the method further comprises the following steps: the glutaraldehyde decamethylammonium bromide solution is used for carrying out a killing experiment on the PRRSV; the killing experiment comprises the following steps:
diluting the glutaraldehyde decamethyl ammonium bromide solution by 100, 500 and 1000 times respectively, mixing the glutaraldehyde decamethyl ammonium bromide solution with a virus suspension according to the proportion of 9, acting for 10, 30 and 60min respectively, then mixing the glutaraldehyde decamethyl ammonium bromide solution with a cell maintenance solution according to the proportion of 1;
inoculating each sample of the glutaraldehyde decamethylammonium bromide solution into 4 wells, each well being 0.1mL, using a negative control of adding only 0.1mL of cell maintenance fluid to Marc-145 and BHK-21 cells;
diluting PRRSV and PRV virus suspensions by 10-fold gradient with a cell maintenance solution, and then sequentially and correspondingly inoculating the virus suspensions with each dilution into a cell culture plate growing into a monolayer cell, namely inoculating PRRSV mixed liquor into Marc-145 cells and inoculating PRV mixed liquor into BHK-21 cells; 4 wells per sample, 0.1mL per well;
placing the inoculated plates at 37 ℃ with 5% CO 2 The incubator is kept still and adsorbed for 1h, residual virus sample liquid is discarded after the virus is adsorbed in cells, 0.2mL of fresh cell maintenance liquid is added, and CO is put in 2 Continuously culturing in incubator for daily useAnd (4) observing the growth condition of the cells by an inverted microscope for 5-7 days continuously, counting and recording when the cells to be subjected to mutation do not develop mutation and the control cells are intact, and obtaining an experimental result.
Preferably, the glutaraldehyde decamethylammonium bromide solution is used for 100% killing of escherichia coli, staphylococcus aureus, candida albicans and bacillus subtilis at-35-0 ℃.
Preferably, the glutaraldehyde decamethylammonium bromide solution is used for 100% killing of pathogenic porcine reproductive and respiratory syndrome virus and pseudorabies virus; the maximum dilution multiple of the glutaraldehyde decamethylammonium bromide solution for 100% killing of the pathogenic porcine reproductive and respiratory syndrome virus is 1; the maximum dilution factor of the glutaraldehyde decamethylammonium bromide solution for 100% killing of the pseudorabies virus in water is 1.
Preferably, the glutaraldehyde decamethyl ammonium bromide solution is used in a wagon balance and periphery of a pigsty with a dilution factor of 1:2000, dilution factor 1 for use in the pig farm and in the vehicle.
According to the specific embodiment provided by the invention, the invention discloses the following technical effects:
the glutaraldehyde decamethylammonium bromide solution has no pungent smell, high stability, long-term storage and obvious sterilization and disinfection effects under the low-temperature condition of (-35-0 ℃); can kill Escherichia coli, staphylococcus aureus, candida albicans, bacillus subtilis, pathogenic blue ear virus, foot and mouth disease virus and H5N1 subtype avian influenza virus 100% at proper dilution. Is suitable for disinfection of farms, public places, equipment, instruments, hatching eggs and the like. The invention mainly has the following characteristics:
(1) The glutaraldehyde decamethylammonium bromide solution has high stability, does not generate layering phenomenon after long-term storage, and can keep high-efficiency sterilization and disinfection effects.
(2) The glutaraldehyde decamethylammonium bromide solution can kill Escherichia coli, staphylococcus aureus, candida albicans, bacillus subtilis, pathogenic blue ear virus and foot and mouth disease virus of pigs by 100 percent even at low temperature and under proper dilution times.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Reference herein to "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment can be included in at least one embodiment of the application. The appearances of the phrase in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments. It is explicitly and implicitly understood by one skilled in the art that the embodiments described herein can be combined with other embodiments.
The terms "comprising" and "having," and any variations thereof, in the description and claims of this application are intended to cover non-exclusive inclusions. For example, the inclusion of a list of steps, processes, methods, etc. is not limited to only those steps recited, but may alternatively include additional steps not recited, or may alternatively include additional steps inherent to such processes, methods, articles, or devices.
In the following examples, glutaraldehyde, ammonium decamethyl bromide, higher fatty alcohol polyoxyethylene ether, and glycerol are commercially available.
The present invention will be described in further detail with reference to specific embodiments in order to make the above objects, features and advantages more apparent and understandable.
Example 1: the glutaraldehyde decamethyl ammonium bromide solution is prepared from the following raw materials in percentage by weight: 6% of glutaraldehyde, 6% of decamethylammonium bromide, 0.3% of high-carbon fatty alcohol polyoxyethylene ether, 15% of glycerol and the balance of deionized water.
The preparation method of the glutaraldehyde-decamethylammonium bromide solution comprises the following steps:
s1, weighing: weighing the raw material components according to the weight percentage of the raw material components of the glutaraldehyde decamethylammonium bromide solution;
s2, preparation of a mixture A: mixing glutaraldehyde and decamethylammonium bromide, and stirring uniformly to obtain a mixture A;
s3, preparation of a mixture B: adding fatty alcohol-polyoxyethylene ether and glycerol into 60ml of deionized water, then adding the mixture A, and stirring and uniformly mixing to obtain a mixture B;
s4, preparing a glutaraldehyde decamethyl ammonium bromide solution: and (3) adjusting the pH value of the mixture B obtained in the step (S2) to 6, adding deionized water to a constant volume of 100ml, uniformly mixing, and subpackaging after passing inspection to obtain the glutaraldehyde decamethyl ammonium bromide solution.
Comparative example 1:
in the formulation of the glutaraldehyde decamethylammonium bromide solution, the difference from example 1 is that in comparative example 1, the higher fatty alcohol polyoxyethylene ether is removed, the amount of deionized water is increased, and the rest of the components and the preparation method refer to example 1.
Comparative example 2:
in the glutaraldehyde decamethylammonium bromide solution formulation, the difference from example 1 is that comparative example 2 is made by removing glycerin, and the rest of the components and the preparation method are referred to example 1.
Test example 1: stability test of glutaraldehyde decamethylammonium bromide solution
1. Test materials: the resulting glutaraldehyde decamethylammonium bromide solutions were prepared for example 1 and comparative example 1.
2. The test method comprises the following steps: referring to disinfectant stability test in appendix of Chinese animal pharmacopoeia (2015 edition), the appearance, smell, pH value and content of glutaraldehyde decamethylammonium bromide solution are recorded after being placed for 5 and 10 days under the conditions of high temperature (60 ℃), illumination (4500 LX +/-500 LX) and low-temperature refrigeration (-15 ℃).
3. And (3) test results: see table 1.
TABLE 1 stability test of glutaraldehyde decamethylammonium bromide solutions
As can be seen from Table 1, the glutaraldehyde/decamethylammonium bromide solution prepared in example 1 of the present invention has substantially unchanged appearance, odor, pH, glutaraldehyde and decamethylammonium bromide content and high stability after being left for 10 days under high temperature, light and low temperature conditions. And the glutaraldehyde decamethyl ammonium bromide solution prepared in the comparative example 1 (fatty alcohol-free polyoxyethylene ether) is layered and turbid after being placed for 10 days under the conditions of high temperature and illumination, and the content of active ingredients is obviously reduced. The glutaraldehyde decamethylammonium bromide solution prepared in comparative example 2 (without glycerol) freezes at low temperature.
Test example 2: common germ kill test (carried out at-15 ℃ environment)
1. Test materials: glutaraldehyde decamethylammonium bromide solution prepared in example 1.
2. Test subjects: escherichia coli (ATCC 8099), staphylococcus aureus (ATCC 6538), candida albicans (ATCC 10231), and Bacillus subtilis (ATCC 6633).
3. The test method comprises the following steps: the bactericidal properties were tested with reference to 2.1.11.3.1 in the Disinfection Specification (Ministry of health 2002). 0.1mL of each sample is taken and is dripped on a steel sheet with the diameter of 1.0cm, 20 mu L of bacterial liquid is respectively smeared on each steel sheet after the samples are dried, the steel sheets are respectively placed in a neutralizer tube for 2 min, 5min, 10min and 20min, and a viable count test is carried out after the neutralization for 10 min. The control group was prepared by directly dropping the bacterial liquid on a sterile steel plate, and the other steps were the same as those of the test group. The environmental temperature is 19.5-20 ℃, the bacterial culture temperature is 37 ℃, the result is observed after 48 hours of culture, and the test is repeated three times.
4. And (3) test results: see table 2.
TABLE 2 glutaraldehyde decamethylammonium bromide solution pathogen kill test
As can be seen from Table 2, the glutaraldehyde decamethylammonium bromide solution can effectively kill Escherichia coli, staphylococcus aureus, candida albicans and Bacillus subtilis, and when the dilution multiple of the aqueous solution is 1 (1000-2000), the Escherichia coli, staphylococcus aureus and Candida albicans can be killed by 100%, and when the dilution multiple of the aqueous solution is 1. Therefore, the glutaraldehyde decamethylammonium bromide solution can kill escherichia coli, staphylococcus aureus, candida albicans and bacillus subtilis by 100% in a low-temperature environment (15 ℃ below zero).
Test example 3: killing test of highly pathogenic Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) and pseudorabies Virus (PRV) with glutaraldehyde-decamethylammonium bromide solution (performed at-15 deg.C)
1. Test materials: glutaraldehyde decamethylammonium bromide solution prepared in example 1.
2. Test subjects: highly pathogenic Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), pseudorabies virus (PRV) and monkey kidney cell Marc-145, which are provided by animal epidemic disease prevention and control key open laboratories of Ministry of agriculture.
3. The test method comprises the following steps:
(1) Monolayer cell culture: the well-grown monkey kidney cells (Marc-145) and hamster kidney cells (BHK-21) were made into cell suspensions, 0.1mL of each was placed in a respective 96-well cell culture plate, the culture plate was placed in a 5% CO2 incubator, and after 24 hours, if the cells grew into a good monolayer, they were removed for use.
(2) Glutaraldehyde decamethylammonium bromide solution was tested for PRRSV kill: the dilution of example 1 is 100, 500 and 1000 times, the dilution is mixed with virus suspension according to the proportion of 9 to act for 10, 30 and 60min respectively, then the dilution is mixed with cell maintenance liquid according to the proportion of 1. Each sample was inoculated into 4 wells, 0.1mL per well. A negative control was made by adding only 0.1mL of cell maintenance fluid to Marc-145 and BHK-21 cells. The PRRSV and PRV virus suspension is diluted by 10 times of gradient with cell maintenance liquid, then the virus suspension of each dilution is sequentially and correspondingly inoculated to a cell culture plate growing into a monolayer cell, namely PRRSV mixed liquid is inoculated to Marc-145 cells, and PRV mixed liquid is inoculated to BHK-21 cells. Each sample was inoculated into 4 wells, 0.1mL per well. Placing the inoculated culture plate in an incubator containing 5 percent CO2 at 37 ℃ for standing and adsorbing for 1h, discarding residual virus specimen liquid after the virus is adsorbed in cells, adding 0.2mL of fresh cell maintenance liquid, placing the culture plate in a CO2 incubator for continuous culture, observing the growth condition of the cells by an inverted microscope every day, continuously observing for 5-7 days, counting and recording the result when the cells with the mutation are not developed and the control cells are intact, and obtaining the table 3.
TABLE 3 killing of viruses by glutaraldehyde decamethylammonium bromide solution
As can be seen from table 3, the maximum dilution of the glutaraldehyde decamethylammonium bromide solution of the present invention at 100% to kill the blue ear disease virus (PRRSV) is 1 1000 at-15 ℃, but the killing time cannot be less than 30min at 1. The highest dilution at 100% to kill pseudorabies virus (PRV) was 1.
Test example 4: killing tests (conducted at-15 ℃ C.) with glutaraldehyde decamethylammonium bromide solution on weighbridge, in piggery and on transport vehicles
Before the test, the dung is firstly cleaned, the pigsty and the ground of the wagon balance are washed by tap water, the floor of a hopper of a transport vehicle and the like, the pigsty is disinfected after being dried, and doors and windows are closed during the disinfection. After diluting the sample by 500, 1000, 2000 and 3000 times, the sample is taken after spraying for 15min, and the non-sterile area around the place is taken as a control. For each test point 3 samples were taken in parallel and the microorganisms were sampled by wiping.
And (3) performing gradient dilution on all samples by 10 times by using sterile physiological saline, then inoculating 0.1mL of sterile physiological saline to plate counting agar, placing the culture dish in a constant-temperature incubator at 37 ℃ for culturing for 18-24h, and counting each plate manually when the microbial colonies grow to the degree that the microbial colonies can be seen by naked eyes. Table 4 was obtained.
TABLE 4 Disinfection Effect of glutaraldehyde decamethylammonium bromide solution on weighbridge, in hog house and transport vehicle
As can be seen from Table 4, the optimal dilution factor for the glutaraldehyde decamethylammonium bromide solution of the present invention for a wagon balance at-15 ℃ is 1:2000, optimal dilution factor for piggery and vehicle is 1.
In conclusion, in the invention, the fatty alcohol-polyoxyethylene ether (high-carbon fatty alcohol-polyoxyethylene ether AEO) is a nonionic surfactant, has good performance at a temperature lower than the cloud point of the nonionic surfactant, and meets the requirement of low-temperature hard water resistance. The glutaraldehyde decamethylammonium bromide solution can be uniformly dispersed, the solution is more stable, and test example 1 can prove that the glutaraldehyde decamethylammonium bromide solution prepared by the method has no pungent smell, is still colorless clear liquid after being placed for 10 days under the conditions of high temperature, illumination and low temperature, keeps the content of effective components between 5.2 and 5.8 percent, and has high stability. More importantly, polyoxyethylene fatty alcohol ether is added into the glutaraldehyde to obviously improve the sterilization effect of the glutaraldehyde. It is possible that the polyoxyethylene fatty alcohol ether can reduce the volatilization of the effective components of glutaraldehyde and decamethylammonium bromide, and the mechanism is not clear. Test example 2 it can be confirmed that the sterilizing effect of the comparative example in which polyoxyethylene fatty alcohol ether is removed is reduced with time.
Glycerol is a colorless, sweet, clear, viscous liquid, odorless, and is commonly referred to as glycerol. Glycerol is the earliest organic alcohol used for antifreeze and is replaced by methanol and ethylene glycol for cost reasons. With the more mature process, the price of the glycerol is greatly reduced. In addition, in practical application, the addition amount of the glycerol of 20 percent is equivalent to the antifreezing effect of the ethylene glycol or ethanol of 40 percent, so that the glycerol can replace the ethylene glycol or the ethanol in consideration of environmental protection and practical cost. The invention is matched with glycerol to reduce the freezing point of the disinfectant, and the test examples 2-4 are all carried out at the temperature of-15 ℃, and the test example 2 can prove that the maximum dilution times of 100% of water solutions for killing escherichia coli, staphylococcus aureus and candida albicans are 1. Test example 3 can prove that the maximum dilution multiple of 100% glutaraldehyde decamethylammonium bromide solution for killing pathogenic porcine reproductive and respiratory syndrome virus is 1; the highest dilution factor for 100% killing of pseudorabies virus (PRV) is 1. Test example 4 it can be demonstrated that the optimal dilution factor for weighbridge and piggery periphery is 1:2000, optimal dilution factor for use in the pig farm and in the vehicle is 1.
Example 2: the glutaraldehyde decamethylammonium bromide solution is different from the glutaraldehyde decamethylammonium bromide solution in the embodiment 1 in that the glutaraldehyde decamethylammonium bromide solution is prepared from the following raw materials in percentage by weight: 4% of glutaraldehyde, 4% of decamethylammonium bromide, 0.2% of high-carbon fatty alcohol polyoxyethylene ether, 6% of glycerol and the balance of deionized water. The glutaraldehyde decamethylammonium bromide solution was prepared in a manner different from that of example 1 in that, in step S4, the pH of mixture B was adjusted to 5.4.
Example 3: the difference between the glutaraldehyde decamethylammonium bromide solution and the embodiment 1 is that the glutaraldehyde decamethylammonium bromide solution is prepared from the following raw materials in percentage by weight: 7% of glutaraldehyde, 7% of decamethylammonium bromide, 2% of high-carbon fatty alcohol polyoxyethylene ether, 20% of glycerol and the balance of deionized water. The preparation of a solution of glutaraldehyde decamethylammonium bromide differed from example 1 in that in step S4 the pH of mixture B was adjusted to 6.6.
The embodiments in the present description are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other.
The principle and the embodiment of the present invention are explained by applying specific examples, and the above description of the embodiments is only used to help understanding the method and the core idea of the present invention; meanwhile, for a person skilled in the art, according to the idea of the present invention, the specific embodiments and the application range may be changed. In view of the above, the present disclosure should not be construed as limiting the invention.
Claims (5)
1. A preparation method of a glutaraldehyde decamethylammonium bromide solution is characterized by comprising the following steps: the method comprises the following steps:
s1, weighing: weighing raw materials, wherein the raw material components comprise 4-7% of glutaraldehyde, 4-7% of decamethylammonium bromide, 0.2-2% of high-carbon fatty alcohol polyoxyethylene ether, 6-20% of glycerol and the balance of deionized water;
s2, preparing a mixture A: mixing glutaraldehyde and decamethylammonium bromide, and stirring uniformly to obtain a mixture A;
s3, preparation of a mixture B: adding fatty alcohol-polyoxyethylene ether and glycerol into 60ml of deionized water, then adding the mixture A, and stirring and uniformly mixing to obtain a mixture B;
s4, preparing a glutaraldehyde decamethyl ammonium bromide solution: adjusting the pH value of the mixture B obtained in the step S2 to 5.4-6.6, adding deionized water and fixing the volume to 100ml to obtain a glutaraldehyde decamethylammonium bromide solution;
the glutaraldehyde decamethyl ammonium bromide solution is used for dilution disinfection with different concentrations according to use scenes;
the maximum dilution multiple of the glutaraldehyde decamethylammonium bromide solution for 100% killing of escherichia coli, staphylococcus aureus and candida albicans is 1.
2. The method of claim 1, further comprising: the glutaraldehyde decamethylammonium bromide solution is used for carrying out a killing experiment on the PRRSV; the killing experiment comprises the following steps:
diluting the glutaraldehyde decamethyl ammonium bromide solution by 100, 500 and 1000 times respectively, mixing the glutaraldehyde decamethyl ammonium bromide solution with a virus suspension according to the proportion of 9, acting for 10, 30 and 60min respectively, then mixing the glutaraldehyde decamethyl ammonium bromide solution with a cell maintenance solution according to the proportion of 1;
inoculating 4 wells of each of the samples of glutaraldehyde decamethylammonium bromide solution, 0.1mL per well, with a negative control of adding only 0.1mL of cell maintenance fluid to Marc-145 and BHK-21 cells;
diluting PRRSV and PRV virus suspensions by 10-fold gradient with a cell maintenance solution, and then sequentially and correspondingly inoculating the virus suspensions with each dilution into a cell culture plate growing into a monolayer cell, namely inoculating PRRSV mixed liquor into Marc-145 cells and inoculating PRV mixed liquor into BHK-21 cells; 4 wells per sample, 0.1mL per well;
placing the inoculated plates at 37 ℃ with 5% CO 2 The incubator is kept still for adsorption for 1 hour, residual virus sample liquid is discarded after the virus is adsorbed in the cells, 0.2mL of fresh cell maintenance liquid is added, and CO is placed 2 Continuously culturing in the incubator, observing the growth condition of the cells by using an inverted microscope every day, continuously observing for 5-7 days, counting and recording when the cells to be subjected to mutation do not develop and the control cells are intact, and obtaining the experimental result.
3. The method for preparing the glutaraldehyde decamethylammonium bromide solution according to claim 1, wherein the glutaraldehyde decamethylammonium bromide solution is used for 100% killing of escherichia coli, staphylococcus aureus, candida albicans, and bacillus subtilis at-35 to 0 ℃.
4. The method for preparing the glutaraldehyde decamethylammonium bromide solution according to claim 1, wherein the glutaraldehyde decamethylammonium bromide solution is used for 100% killing of pathogenic porcine reproductive and respiratory syndrome viruses and pseudorabies viruses; the maximum dilution multiple of the glutaraldehyde decamethylammonium bromide solution for 100% killing of the pathogenic porcine reproductive and respiratory syndrome virus is 1; the maximum dilution factor of the glutaraldehyde decamethylammonium bromide solution for 100% killing of the pseudorabies virus in water is 1.
5. The method of claim 1, wherein the glutaraldehyde decamethylammonium bromide solution is diluted by a factor of 1:2000, dilution factor 1.
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