CN114984194A - Composite lysozyme disinfectant - Google Patents

Composite lysozyme disinfectant Download PDF

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Publication number
CN114984194A
CN114984194A CN202210687550.XA CN202210687550A CN114984194A CN 114984194 A CN114984194 A CN 114984194A CN 202210687550 A CN202210687550 A CN 202210687550A CN 114984194 A CN114984194 A CN 114984194A
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disinfectant
lysozyme
extract
compound
compound lysozyme
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谢清华
周东
胡晓飞
董雯
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Shandong Bopeng Biotechnology Co ltd
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Shandong Bopeng Biotechnology Co ltd
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    • A61K47/10—Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
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Abstract

The invention belongs to the technical field of disinfectant preparations, and particularly relates to a composite lysozyme disinfectant, which comprises 1-5 g/L of lauramidopropyl betaine, 10000-200000U/mL of lysozyme, 5-10 g/L of inorganic salt, 1-5 g/L of peptone, 0.1-1 g/L of glycosylated phosphatidylinositol anchoring protein crude extract, 5-10 mL/L of glycerol and 0.2-0.5 g/L of biological preservative; by forming a complex of glycosylated phosphatidylinositol-anchor protein and lysozyme, the glycosylated phosphatidylinositol-anchor protein can be specifically bound to bacterial peptidoglycan to capture pathogenic bacteria, and lysozyme destroys the captured bacterial cell wall, thereby killing bacteria. Meanwhile, the tropism of bacteria is utilized, the peptone component is increased, the aggregation of the bacteria is increased, the germs are killed intensively, and the bactericidal antibacterial cream can be used as a spray, is atomized in the air and can be sprayed on hands and intact skin, and has good bactericidal and bacteriostatic effects.

Description

Composite lysozyme disinfectant
Technical Field
The invention belongs to the technical field of disinfectant preparations, and particularly relates to a compound lysozyme disinfectant.
Background
The prior skin disinfectants are mostly chemical reagents, mainly comprise ethanol, iodine and iodine-containing preparations, chlorhexidine, quaternary ammonium salt compounds, triclosan and the like, wherein ethanol solution with the concentration of 60-80% has the best bactericidal effect but poor persistence and narrow bactericidal spectrum, so the ethanol solution is usually only used as common external disinfectant, the iodine and iodine-containing preparations are also skin disinfectants widely used clinically, but certain color is easy to stain the skin to cause the problem of difficult elution, the chlorhexidine has weak effect on gram-negative bacteria and fungi, the quaternary ammonium salt compounds have the narrow bactericidal spectrum and poor effect on gram-negative bacteria, and the triclosan is an efficient broad-spectrum antibacterial agent but has the bactericidal effect relative to the ethanol, the iodine and iodine-containing preparations, Chlorhexidine and quaternary ammonium salt compounds are weaker.
The large difference in cell wall peptidoglycan content between gram-positive bacteria, whose cell wall consists of up to 40 layers of peptidoglycan and teichoic acid, and gram-negative bacteria, which typically have only a single layer of peptidoglycan without teichoic acid sandwiched between an inner membrane and an outer membrane containing lipopolysaccharides, explains why gram-positive bacteria are typically sensitive to lysozyme, but not gram-negative bacteria.
However, after the lysozyme is simply added with the disinfectant components, the comprehensive disinfection effect of the lysozyme is not ideal, so how to realize the maximum effect of the lysozyme on gram-negative bacteria in the disinfectant is an urgent problem to be solved.
Reference documents: [1] yangzhouke, in China, etc. preliminary study on the bacteriostatic action and mechanism of lysozyme of marine microorganisms, advanced fishery science, 2005,26(005):62-68.
Disclosure of Invention
In order to solve the technical problems, the composite lysozyme disinfectant is provided, a compound is formed by glycosylation phosphatidylinositol anchor protein and lysozyme, the glycosylation phosphatidylinositol anchor protein can be specifically combined with bacterial peptidoglycan to capture pathogenic bacteria, and the lysozyme destroys the cell wall of the captured bacteria, thereby killing the bacteria. Meanwhile, the tropism of bacteria is utilized, the peptone component is increased, the aggregation of the bacteria is increased, the germs are killed intensively, the composite lysozyme disinfectant can be used as a spray, is atomized in the air, and can also be sprayed on hands and intact skin, has good sterilization and bacteriostasis effects, is long in duration and free from toxicity to human bodies, and has a strong sterilization effect on escherichia coli, streptococcus, salmonella enteritidis, candida albicans, staphylococcus aureus, particularly various epidemic diseases such as gram-positive bacteria and the like.
If not stated, the compound lysozyme disinfectant is also called disinfectant for short.
Specifically, the technical scheme of the invention is as follows:
1. a compound lysozyme disinfectant comprises the following components in percentage by weight:
Figure BDA0003700222410000021
the inorganic salt is at least one of sodium chloride, potassium chloride, calcium chloride, sodium sulfate and sodium bicarbonate;
the biological preservative is at least one of nisin, natamycin and polylysine.
The invention also comprises a preparation method of the glycosylated phosphatidylinositol anchor protein crude extract, which specifically comprises the following steps: designing a glycosylated phosphatidylinositol-anchored protein primer, amplifying by taking a rice cDNA library plasmid DNA as a template, recovering a product, connecting with a pMD19-T vector, transforming escherichia coli DH5 alpha competent cells, extracting plasmids, carrying out enzyme digestion and connection with a plant expression empty vector pCB, constructing a glycosylated phosphatidylinositol-anchored protein expression vector, electrically shocking and transforming agrobacterium GV3101, coating the transformed agrobacterium GV on an SOB solid culture medium containing three resistances, inverting and culturing for 2d at 28 ℃, selecting a colony PCR (polymerase chain reaction) to identify correct single spots to prepare an infection solution, transfecting the plant rice in a mode, culturing for 1 week, reducing root systems, grinding, and filtering by filter paper to obtain a crude extract.
Further, the pH value of the compound lysozyme disinfectant is 6-8.
The invention also comprises a preparation method of the compound lysozyme disinfectant, which comprises the following steps:
mixing lauramidopropyl betaine, lysozyme, inorganic salt, peptone, glycosylated phosphatidylinositol anchor protein crude extract, glycerol and a biological preservative in water at the temperature of 5-40 ℃ and stirring for 1-12 hours to obtain a finished product.
Further, the compound lysozyme disinfectant also comprises a plant extract, wherein the plant extract is at least one of a green tea extract, an aloe extract, a centella asiatica extract, a lavender extract, a lemon extract, a peach extract, a sage extract, a ginkgo biloba leaf extract and a honeysuckle flower extract; the plant extract has the effect of improving the smell of the disinfectant, and accounts for 0.5 to 1 percent of the weight of the disinfectant.
The invention also provides application of the compound lysozyme disinfectant in air disinfection/skin external disinfection.
Compared with the prior art, the invention has the following beneficial effects:
1) the specificity is strong: according to the invention, through the characteristics of glycosylated phosphatidylinositol anchor protein, the bacterial polysaccharide component can be specifically combined, various pathogenic bacteria can be specifically captured, and lysozyme is combined for killing;
2) the bactericidal effect is strong: the invention combines lysozyme and glycosylation phosphatidylinositol anchoring protein, the lysozyme destroys the bacterial cell wall, meanwhile, the bacterium is gathered by increasing the nutrient content of peptone and utilizing the characteristic that the bacterium tends to the nutrient content, the bacterium can be intensively killed, and the double effects are achieved, thereby enhancing the sterilization effect. Due to the combination of glycosylated phosphatidylinositol anchoring protein, the killing probability to gram-negative bacteria is increased, and the defect that lysozyme is not sensitive to gram-negative bacteria is overcome;
3) the stability is good: according to the invention, the disinfection inert component lauramidopropyl betaine, glycerol and inorganic salt form a stable group, and a protective layer is formed on the surfaces of lysozyme and glycosylated phosphatidylinositol anchoring protein, so that the interference caused by the external environment is reduced;
4) safety and environmental protection: compared with chemical disinfectants, the invention adopts biological disinfection components and plant crude extract, and is safe, nontoxic and free of side effect.
Detailed Description
The invention is described below by means of specific embodiments. Unless otherwise specified, the technical means used in the present invention are well known to those skilled in the art. In addition, the embodiments should be considered illustrative, and not restrictive, of the scope of the invention, which is defined solely by the claims. It will be apparent to those skilled in the art that various changes or modifications in the components and amounts of the materials used in these embodiments can be made without departing from the spirit and scope of the invention.
The preferred compound lysozyme disinfectant of the invention comprises the following components by weight:
Figure BDA0003700222410000041
Figure BDA0003700222410000051
the lauramidopropyl betaine may be present in an amount of 1g/L, 1.5g/L, 1.8g/L, 2g/L, 3g/L, 4g/L or 5g/L, but is not limited to the recited values, and other values not recited in the recited range of values are equally applicable; the content of lysozyme can be 10000U/mL, 12000U/mL, 15000U/mL, 16000U/mL or 20000U/mL, etc., but is not limited to the recited values, and other unrecited values within the listed values are also applicable; the inorganic salt can be present in an amount of 5g/L, 5.5g/L, 6g/L, 6.5g/L, 7g/L, 7.5g/L, 8g/L, 8.5g/L, 9g/L, or 10g/L, and the like, but is not limited to the recited values, and other non-recited values within the recited range are equally applicable; the content of peptone may be 1g/L, 2g/L, 3g/L, 4g/L or 5g/L, etc., but is not limited to the enumerated values, and other unrecited values within the enumerated values are also applicable; the content of the crude extract of glycosylated phosphatidylinositol anchor protein may be 0.1g/L, 0.2g/L, 0.3g/L, 0.4g/L, 0.5g/L, 0.6g/L, 0.7g/L, 0.8g/L, 0.9g/L, or 1g/L, etc., but is not limited to the recited values, and other values not recited in the listed values are also applicable; the glycerol content can be 5mL/L, 6mL/L, 7mL/L, 8mL/L, 9mL/L, or 10mL/L, etc., but is not limited to the recited values, and other unrecited values within the recited range of values are equally applicable; the content of the biological preservative can be 0.2g/L, 0.3g/L, 0.4g/L or 0.5g/L, etc., but is not limited to the recited values, and other values not recited within the listed value range are also applicable.
The inorganic salts of the present invention also function as solubilizers, and other solubilizing inorganic salts are within the choice of the present invention.
Preferably, the compound lysozyme disinfectant comprises the following components in parts by weight:
Figure BDA0003700222410000052
Figure BDA0003700222410000061
preferably, the compound lysozyme disinfectant comprises the following components in percentage by weight:
Figure BDA0003700222410000062
the amount of each component is not limited to the specific value, and the ratio of the corresponding components is within the protection scope of the present invention.
Some preferred embodiments of the present invention further comprise a plant extract, wherein the plant extract is one or a combination of green tea extract, aloe extract, centella asiatica extract, lavender extract, lemon extract, peach extract, sage extract, ginkgo biloba extract and honeysuckle extract; for example, a mixture of green tea extract and aloe extract, a mixture of aloe extract, centella extract and lavender extract, a mixture of lemon extract and peach extract, a mixture of sage extract, ginkgo biloba extract and honeysuckle extract, but not limited to the listed combinations, and other non-listed single substances or combinations are equally suitable.
In the embodiment of the invention, lauramidopropyl betaine, lysozyme, inorganic salt, peptone, glycosylated phosphatidylinositol anchoring protein crude extract, glycerol and a biological preservative are mixed in water at 5-40 ℃ for 1-12 hours to obtain the finished product of the composite lysozyme disinfectant.
In the embodiment of the present invention, the mixing is performed by stirring, and the stirring time may be 3 hours, 4 hours, 5 hours, 6 hours, or the like, but is not limited to the values listed, and other values not listed in the range of the values are also applicable.
The embodiment of the invention also provides application of the compound lysozyme disinfectant.
It should be noted that, when the composite lysozyme disinfectant provided in the embodiment of the present invention is applied, a user may adjust the usage amount and the usage concentration thereof according to actual needs, and may directly use the stock solution of the disinfectant, or may dilute the stock solution of the disinfectant.
The broad-spectrum antiviral and antibacterial disinfectant provided by the embodiment of the invention can be directly sprayed on the surface of an object to be sterilized, and can also be directly acted on the skin of the hand of a human body, and the application forms of the broad-spectrum antiviral and antibacterial disinfectant include but are not limited to spraying of a spray, wiping of a carrier, smearing of a hand sanitizer and the like.
The present invention will be further described with reference to specific examples. The experimental methods used in the examples of the present invention are all conventional methods unless otherwise specified; materials, reagents and the like used in examples of the present invention are commercially available unless otherwise specified.
Example 1
The preparation of the crude extract of glycosylated phosphatidylinositol anchor protein comprises the following steps:
designing a glycosylated phosphatidylinositol anchor protein primer, taking rice cDNA library plasmid DNA as a template for amplification, recovering a product, connecting the product with a pMD19-T vector, transforming escherichia coli DH5 alpha competent cells, extracting plasmids, carrying out enzyme digestion to connect a plant expression empty vector pCB, constructing a glycosylated phosphatidylinositol anchor protein expression vector, electrically shocking and transforming agrobacterium GV3101, coating the transformed agrobacterium GV3101 on a SOB solid culture medium containing three resistances, carrying out inversion dark culture at 28 ℃ for 2d, selecting a bacterial colony and identifying correct single spots by PCR to prepare an infection solution, transfecting the plant rice in a mode, culturing for 1 week, reducing root systems, grinding, and filtering through filter paper to obtain a crude extract.
The composite lysozyme disinfectant comprises the following components in percentage by weight:
Figure BDA0003700222410000081
dissolving the above components in 1L water in sequence of sodium chloride, lauramidopropyl betaine, glycosylated phosphatidylinositol anchor protein crude extract, lysozyme, glycerol, peptone, nisin, and fructus Citri Limoniae extract, and mixing and stirring at room temperature for 2h to obtain the final product.
The pH value of the compound lysozyme disinfectant is 6.9.
Example 2
The procedure for the preparation of a crude extract of glycosylated phosphatidylinositol-anchored protein is the same as in example 1.
The embodiment provides a compound lysozyme disinfectant, which comprises the following components in concentration: 10g/L potassium chloride, 1g/L lauramidopropyl betaine, 1g/L glycosylated phosphatidylinositol anchor protein crude extract, 20000U/mL lysozyme, 5mL/L glycerol, 5g/L peptone, 0.5g/L polylysine and 1% (weight) of green tea extract are dissolved in 1L water according to the sequence and content concentration, and the mixture is mixed and stirred for 2 hours at normal temperature to obtain the finished product. The pH value of the compound lysozyme disinfectant is 6.3.
Example 3
The procedure for the preparation of a crude extract of glycosylated phosphatidylinositol-anchored protein is the same as in example 1.
The embodiment provides a compound lysozyme disinfectant, which comprises the following components in concentration: 5g/L sodium sulfate, 1g/L lauramidopropyl betaine, 1g/L glycosylated phosphatidylinositol anchor protein crude extract, 200000U/mL lysozyme, 5mL/L glycerol, 5g/L peptone, 0.5g/L natamycin and 1% (weight) lavender extract are dissolved in 1L water according to the sequence and content concentration, and are mixed and stirred for 2 hours at normal temperature to obtain a finished product.
The pH value of the compound lysozyme disinfectant is 7.2.
Comparative example 1
The comparative example provides a compound lysozyme disinfectant, which has the same composition and content as those of the example 1 except that the compound lysozyme disinfectant does not contain the glycosylated phosphatidylinositol anchor protein crude extract, and the preparation method is also the same as the example 1.
Comparative example 2
The comparative example provides a compound lysozyme disinfectant, which has the same components and content as those of the disinfectant in the example 1 except that no peptone is contained, and the preparation method is also the same as that of the disinfectant in the example 1.
Test of Sterilization Effect
The sterilizing effects of the sterilizing solutions prepared in examples 1 to 3 and comparative examples 1 to 2 on escherichia coli were tested, and the test results are shown in table 1.
TABLE 1 Performance test of disinfectant solutions prepared in examples 1 to 3 and comparative examples 1 to 2
Figure BDA0003700222410000091
Figure BDA0003700222410000101
In order to further verify the sterilizing capability of the disinfectant provided by the invention on various bacteria, the compound lysozyme disinfectant prepared in the embodiment 1 is selected to test various microorganisms to obtain test results shown in tables 2 to 5, wherein the concentration x% given in tables 2 to 5 is calculated according to the original concentration of the antibacterial disinfectant, and the test conditions are as follows: the results were obtained in a standardized manner with low albumin content (0.03%)/dirty with high albumin content (0.3%).
TABLE 2 Bactericidal Effect against Salmonella enteritidis
Figure BDA0003700222410000102
TABLE 3 fungicidal Effect against Candida albicans
Figure BDA0003700222410000103
Note: the bacterial test standard is EN 1276.
TABLE 4 Bactericidal Effect against Staphylococcus aureus
Figure BDA0003700222410000104
Note: the bacterial test standard is EN 1276.
TABLE 5 bactericidal Effect on Streptococcus
Figure BDA0003700222410000105
As can be seen from tables 1-5, the composite lysozyme disinfectant provided by the invention has good sterilization and bacteriostasis effects, is long in duration, can play a role in continuous sterilization and bacteriostasis, and has a strong sterilization effect on various epidemic infectious diseases such as escherichia coli, streptococcus, salmonella enteritidis, candida albicans, staphylococcus aureus and the like.

Claims (10)

1. The compound lysozyme disinfectant comprises the following components in parts by weight:
Figure FDA0003700222400000011
the inorganic salt is at least one of sodium chloride, potassium chloride, calcium chloride, sodium sulfate and sodium bicarbonate;
the biological preservative is at least one of nisin, natamycin and polylysine.
2. The compound lysozyme disinfectant of claim 1, which comprises the following components in percentage by weight:
Figure FDA0003700222400000012
the inorganic salt is sodium chloride;
the biological preservative is at least one of nisin, natamycin and polylysine.
3. The compound lysozyme disinfectant of claim 1, which comprises the following components in percentage by weight:
Figure FDA0003700222400000021
4. the compound lysozyme disinfectant of claim 1, which comprises the following components in percentage by weight:
Figure FDA0003700222400000022
5. the compound lysozyme disinfectant of claim 1, which comprises the following components in percentage by weight:
Figure FDA0003700222400000023
Figure FDA0003700222400000031
6. the compound lysozyme disinfectant of any one of claims 1 to 5, wherein the preparation method of the glycosylated phosphatidylinositol anchor protein crude extract comprises the following steps: constructing a glycosylated phosphatidylinositol-anchored protein expression vector, culturing for 1 week after transfecting a model plant rice, then reducing root systems, grinding, and filtering by filter paper to obtain a crude extract.
7. The compound lysozyme disinfection solution of any one of claims 1 to 5, wherein the pH of the compound lysozyme disinfection solution is 6 to 8.
8. The compound lysozyme disinfectant of any one of claims 1 to 5, wherein the preparation method of the compound lysozyme disinfectant comprises the following steps:
and mixing lauramidopropyl betaine, lysozyme, inorganic salt, peptone, glycosylated phosphatidylinositol anchor protein crude extract, glycerol and a biological preservative in water at the temperature of 5-40 ℃ for 1-12 hours to obtain the compound lysozyme disinfectant.
9. The compound lysozyme disinfectant of any one of claims 1 to 5, further comprising a plant extract, wherein the plant extract is at least one of a green tea extract, an aloe extract, a centella asiatica extract, a lavender extract, a lemon extract, a peach extract, a sage extract, a ginkgo biloba extract and a honeysuckle extract; the plant extract accounts for 0.5-1% (by weight) of the composite lysozyme disinfectant.
10. The use of the compound lysozyme disinfectant of claim 1 in air disinfection/skin external disinfection.
CN202210687550.XA 2022-06-17 2022-06-17 Composite lysozyme disinfectant Pending CN114984194A (en)

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CN101461379A (en) * 2007-12-18 2009-06-24 李萍 Composite lysozyme disinfecting liquid
CN104800095A (en) * 2015-04-22 2015-07-29 江苏雪豹日化有限公司 Biological lysozyme compound preparation and preparation method thereof
CN105211128A (en) * 2015-10-13 2016-01-06 蒋爱英 A kind of clinical laboratory Chinese medicine special multienzyme compound disinfectant and preparation method thereof
CN113967251A (en) * 2021-10-09 2022-01-25 山东凯普润消毒灭菌技术有限公司 Bacteriostatic agent, hand sanitizer containing bacteriostatic agent and preparation method of hand sanitizer
CN114159337A (en) * 2021-11-02 2022-03-11 义乌市欧雅化妆品有限公司 Facial antibacterial care solution containing recombinant human lysozyme

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Publication number Priority date Publication date Assignee Title
CN101461379A (en) * 2007-12-18 2009-06-24 李萍 Composite lysozyme disinfecting liquid
CN104800095A (en) * 2015-04-22 2015-07-29 江苏雪豹日化有限公司 Biological lysozyme compound preparation and preparation method thereof
CN105211128A (en) * 2015-10-13 2016-01-06 蒋爱英 A kind of clinical laboratory Chinese medicine special multienzyme compound disinfectant and preparation method thereof
CN113967251A (en) * 2021-10-09 2022-01-25 山东凯普润消毒灭菌技术有限公司 Bacteriostatic agent, hand sanitizer containing bacteriostatic agent and preparation method of hand sanitizer
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116492268A (en) * 2023-06-21 2023-07-28 潍坊凯利来生物科技有限公司 Hand washing disinfection effervescent tablet and preparation method thereof
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