CN112716879A - Nine-ingredient anti-plague hand sanitizer and preparation method thereof - Google Patents

Abstract

The invention discloses nine-ingredient anti-plague hand sanitizer and a preparation method thereof, and belongs to the technical field of prevention and treatment of epidemic diseases. The hand sanitizer is prepared from the following raw materials: 140-160 parts of fructus chebulae immaturus, 140-160 parts of aconitum pendulum, 140-160 parts of calamus, 140-160 parts of black sulfur, 140-160 parts of mukul myrrh, 140-160 parts of kyo ink, 4-6 parts of asafetida, 0.7-1.2 parts of musk and 65-75 parts of calculus bovis factitius. The nine-ingredient plague-preventing hand sanitizer disclosed by the invention has a good bacteriostatic effect and can effectively prevent plague.

Description

Nine-ingredient anti-plague hand sanitizer and preparation method thereof

Technical Field

The invention relates to nine-ingredient anti-plague hand sanitizer and a preparation method thereof, and belongs to the technical field of prevention and treatment of epidemic diseases.

Background

Plague is an infectious disease caused by some strongly pathogenic substances, such as bacteria and viruses; generally, the environmental sanitation is not good after natural disasters; the epidemic disease comprises the following steps: black death, plague, smallpox, influenza, rabies, tuberculosis, dengue fever, aids, ebola virus, etc. Wu in Ming Dynasty can witness the tragic state of epidemic and death at that time, and find that pestilence is mainly caused by 'entering the mouth and nose' or mutual contact; it is toxic and drastic, unlike six excesses; epidemic prevention measures such as isolation, disinfection and the like are adopted in combination with the characteristics of the disease attack; the main treatment is to clear pestilence and detoxify toxicity. Measles and other respiratory tract infectious diseases are caused by 'between breaths and exogenous pathogens', and intestinal infectious diseases such as cholera and dysentery are also found to be caused by improper eating or rotten food; the skin infectious diseases are caused by contacting with the infection of insect poison and wind evil, and the prevention and treatment effects of traditional Chinese medicines on some serious infectious diseases are very obvious in recent decades.

The traditional Chinese and Tibetan medicine has the main characteristics of preventing pestilence: effective measures are taken before diseases occur, so that the diseases are prevented, and the traditional Chinese medicine, the western medicine or other medicines are consistent; once the disease is developed, effective measures are taken in time to prevent the development, transmission or recurrence of the disease, which is the key point for traditional Chinese medicine prophylactics to distinguish from other preventive medicine. Because the plague is mainly caused by the mouth and the nose, the cleaning treatment of the skin exposed to the air such as the hands and the like is very important.

Disclosure of Invention

The invention aims to provide a nine-ingredient anti-plague hand sanitizer, which is prepared from the following raw materials: 140-160 parts of fructus chebulae immaturus, 140-160 parts of aconitum pendulum, 140-160 parts of calamus, 140-160 parts of black sulfur, 140-160 parts of mukul myrrh, 140-160 parts of kyo ink, 4-6 parts of asafetida, 0.7-1.2 parts of musk and 65-75 parts of calculus bovis factitius.

Preferably, the hand sanitizer is prepared from the following raw materials: 150 parts of fructus chebulae immaturus, 150 parts of aconitum pendulum, 150 parts of acorus calamus, 150 parts of black sulfur, 150 parts of Mokul myrrh, 150 parts of Jingmo, 5 parts of asafetida, 1 part of musk and 70 parts of calculus bovis factitious.

The invention also aims to provide a preparation method of the nine-ingredient anti-plague hand sanitizer, which specifically comprises the following steps:

weighing fructus Chebulae Immaturus, radix Aconiti Szechenyiani, rhizoma Acori Calami, Sulfur, Myrrha, JINGJIU, resina Ferulae, Moschus, and artificial calculus bovis respectively to obtain mixed Chinese medicinal materials, and mixing the materials according to the ratio of medicinal materials-anhydrous ethanol: mixing the raw materials at a ratio of 1:30, refluxing for three times at 50 ℃, combining the supernatants of the three times, concentrating, drying and uniformly mixing to obtain a concentrated product with the crude drug content of 400-500 g/L, adding conventional auxiliary materials of the hand sanitizer into the concentrated product, and preparing the nine-ingredient anti-plague hand sanitizer according to a conventional method.

The invention has the beneficial effects that:

(1) the compatible formula of the anti-plague hand sanitizer purely belongs to natural Tibetan medicinal materials, is beneficial to human bodies and harmless, and has no stimulation and toxic or side effect on skin proved by experiments.

(2) The anti-plague liquid soap has good inhibition activity on the fusion of new coronavirus ACE2 target spots by the components in the anti-plague liquid soap through experiments, can seal over 95 percent of ACE2 receptors in the experimental state, has an inhibition effect on infected cells of the new coronavirus, and also has an antibacterial effect on common bacteria and fungi, so the anti-plague liquid soap has the effects of avoiding plague and preventing plague.

Drawings

FIG. 1 shows the results of affinity detection of binding between blank S protein and ACE2 receptor on CM5 chip;

FIG. 2 shows the result of detecting the binding affinity of blank group S protein to ACE2 receptor on CM5 chip under the action of negative control drug;

FIG. 3 shows the result of affinity detection of coupling between blank group S protein and ACE2 receptor of CM5 chip under the action of positive control drug;

FIG. 4 shows the result of affinity detection of coupling between blank group S protein and ACE2 receptor of CM5 chip under the action of nine anti-pestilence melanophores.

Detailed Description

The present invention is further described in detail with reference to the following specific examples, but the scope of the present invention is not limited to the above description.

Example 1

Nine-ingredient anti-plague liquid soap is prepared from the following raw materials: 150g of fructus chebulae immaturus, 150g of aconitum pendulum, 150g of acorus calamus, 150g of black sulfur, 150g of mukul myrrh, 150g of Beijing ink, 5g of asafetida, 1g of musk and 70g of calculus bovis factitius.

The nine-ingredient plague-preventing hand sanitizer described in this example is prepared by the following method: weighing fructus Chebulae Immaturus, radix Aconiti Szechenyiani, rhizoma Acori Calami, Sulfur, Myrrha, JINGJIU, resina Ferulae, Moschus, and artificial calculus bovis respectively to obtain mixed Chinese medicinal materials, and mixing the materials according to the ratio of medicinal materials-anhydrous ethanol: mixing at a ratio of 1:30, refluxing for three times at 50 ℃, combining the three supernatants, concentrating, drying, and mixing uniformly to obtain a concentrated product with the crude drug content of 450g/L, adding conventional adjuvants of the hand sanitizer into the concentrated product, and preparing the nine-ingredient anti-plague hand sanitizer according to a conventional method.

Example 2

Nine-ingredient anti-plague liquid soap is prepared from the following raw materials: 160g of fructus chebulae immaturus, 145g of aconitum pendulum, 140g of acorus calamus, 160g of black sulfur, 160g of mukul myrrh, 140g of Beijing ink, 4g of asafetida, 0.7g of musk and 65g of calculus bovis factitius.

The nine-ingredient plague-preventing hand sanitizer described in this example is prepared by the following method: weighing fructus Chebulae Immaturus, radix Aconiti Szechenyiani, rhizoma Acori Calami, Sulfur, Myrrha, JINGJIU, resina Ferulae, Moschus, and artificial calculus bovis respectively to obtain mixed Chinese medicinal materials, and mixing the materials according to the ratio of medicinal materials-anhydrous ethanol: mixing at a ratio of 1:30, refluxing for three times at 50 ℃, combining the three supernatants, concentrating, drying, and mixing uniformly to obtain a concentrated product with the crude drug content of 500g/L, adding conventional adjuvants of the hand sanitizer into the concentrated product, and preparing the nine-ingredient anti-plague hand sanitizer according to a conventional method.

Example 3

Nine-ingredient anti-plague liquid soap is prepared from the following raw materials: 140g of fructus chebulae immaturus, 160g of aconitum pendulum, 160g of acorus calamus, 145g of black sulfur, 140g of mukul myrrh, 160g of Beijing ink, 6g of asafetida, 1.2g of musk and 75g of calculus bovis factitius.

The nine-ingredient plague-preventing hand sanitizer described in this example is prepared by the following method: weighing fructus Chebulae Immaturus, radix Aconiti Szechenyiani, rhizoma Acori Calami, Sulfur, Myrrha, JINGJIU, resina Ferulae, Moschus, and artificial calculus bovis respectively to obtain mixed Chinese medicinal materials, and mixing the materials according to the ratio of medicinal materials-anhydrous ethanol: mixing at a ratio of 1:30, refluxing for three times at 50 ℃, combining the three supernatants, concentrating, drying, and mixing uniformly to obtain a concentrated product with the crude drug content of 400g/L, adding conventional adjuvants of the hand sanitizer into the concentrated product, and preparing the nine-ingredient anti-plague hand sanitizer according to a conventional method.

In order to further verify the effect of the nine-ingredient plague-preventing hand sanitizer of the present invention, a series of experiments were performed as follows:

first, the anti-neocoronary pneumonia membrane fusion activity of the Chinese medicinal formula described in this example was determined using Biacore technology.

(1) Experimental Material

Experimental proteins: 293F human embryo kidney cell expression system novel coronavirus spike protein S1 subunit, 293F human embryo kidney cell expression system human ACE2 extramembranous binding region.

(2) Experimental raw materials and reagents

Raw materials: BIAcore Series S Sensor Chip CM5 Sensor Chip and EDC/NHS coupling reagent were purchased from GE Healthcare, 0.22 filter membrane from Millibo China Co., Ltd, 10XPBS buffer from Beijing Soilebao; conventional reagents: sodium chloride, Tween, dimethyl sulfoxide, formic acid, and isopropanol were obtained from Aladdin reagent.

(3) Selection of SPR experiment buffer solution and sample preparation

Total system buffer: measuring deionized water filtered by an 800mL filter by using a measuring cylinder with the volume of 1000mL, measuring 102mL 10XPBS buffer solution by using a measuring cylinder with the volume of 250mL, adding the buffer solution into the 1000mL measuring cylinder to be diluted and uniformly mixed with the deionized water, transferring the diluted deionized water into a 1000mL glass bottle, adding a magnetic rotor, placing the mixed solution on a magnetic stirrer to be uniformly stirred, adjusting the pH value of the mixed solution to 7.4, transferring the mixed solution into a measuring cylinder with the volume of 1000mL, metering the volume of the filtered deionized water to 1000mL and uniformly mixing the deionized water, and finally performing negative pressure filtration by using a 0.22-aperture filter membrane filter to obtain 1.02X PBS buffer solution.

Preparing a protein sample: and (3) discharging the partitioned Covid-19spike and ACE2 protein from a refrigerator at the temperature of-80 ℃, then diluting by using a whole system buffer solution, finally diluting to obtain a Covid-19spike and ACE2 protein solution of 100 mu g/mL, and storing in a refrigerator at the temperature of 4 ℃ for later use.

(4) Chip processing and coupling

The sample compartment temperature was set at 25 ℃. The specific detection parameters of ACE2 protein coupling are: channel 1 is a reference channel and is only blocked by ethanolamine; the channel 2 is a detection channel, the EDC/NHS activation time is set to be 70s, the ACE2 protein contact time is 120s, the ethanolamine blocking time is 30s, and the flow rate is set to be 2 mu L/min; the experimental parameters of the regeneration reagent are as follows: the injection time is set to be 30s, and the flow rate is set to be 30 mu L/min; and finally, analyzing and processing the data by using BIAcore professional software.

(5) ACE2 butt coupling experiment

Preparing a buffer solution: the whole system is a buffer solution.

Chip processing: placing the CM5 biosensor chip in BIAcore X100, and simultaneously using a whole system buffer solution as a flow phase to perform washing regulation on a machine; A0.01M aqueous solution of sodium hydroxide was used as a regenerant, and the regenerant was thoroughly shaken and mixed by a shaker for use.

The experimental conditions are as follows: the temperature of the biosensor chip of BIAcore X100 was set to 25 ℃; the specific detection method for detecting the affinity comprises the following steps: 1. preparing S protein with different concentrations, gradually diluting by 5 times from 0.1 mug/mL, and preparing 5 coupling samples with different concentrations; the samples were placed in order in the sample chamber and the control and regeneration fluids were placed, operating according to the procedure of the affinity program, for one cycle of analysis.

Competitive assay of the Chinese medicinal formulation described in this example

Preparing a buffer solution: the whole system is a buffer solution.

Preparing multi-concentration S protein: setting the initial experimental concentration of the S protein to be 0.1 mu g/mL by taking the KD value of the multi-concentration detection data as a reference; in view of the requirement of the experiment, the Control solution is required to be PBS, and the S protein solution with different concentrations and the sample to be tested are required to be mixed in advance.

Chip processing: placing the CM5 biosensor chip in BIAcore X100, and simultaneously using a whole system buffer solution as a flow phase to perform washing regulation on a machine; after 15 seconds of treatment with the regenerating solution, the chip surface was allowed to equilibrate with the whole system buffer for 7 minutes.

The experimental conditions are as follows: the temperature of the biosensor chip of BIAcore X100 was set to 25 ℃; the specific detection method for detecting the affinity comprises the following steps: preparing S protein with different concentrations, gradually diluting by 5 times from 0.1 mug/mL, preparing 5 coupling samples with different concentrations, and premixing the skin bacteriostatic solution with the concentration of 0.1mg/mL and the S protein solution with different concentrations. The samples were placed in order in the sample chamber and the control and regeneration fluids were placed, operating according to the procedure of the affinity program, for one cycle of analysis.

And (3) test results:

(1) 10000pg/mL, 2000pg/mL, 400pg/mL, 80pg/mL and 16pg/mL spike protein solutions are prepared respectively and are placed in the sample bin No. 2-6 according to the sequence, the blank whole system buffer solution is placed in the bin No. 1, and the regeneration solution is placed in the bin No. 7. The Affinity program in BIAcore X100 was started and one cycle of Affinity assays was started.

The results are shown in fig. 1, 5S proteins with different concentrations are respectively coupled with the chip, the coupling amount of the S protein with different concentrations can be used for measuring the binding parameters of the S protein and the ACE2 receptor in a blank state by a fitting mode, and the analysis results are shown in table 1.

TABLE 1

KD(M)	SE(KD)	Rmax(RU)	SE(Rmax)	offset(RU)	SE(offset)
						7.558E-7	2.5E-7
		1059.9	2.9E+2	19.3	1.6

The binding constant KD (M) of the S protein and the ACE2 receptor in a blank state is as high as 7.558E-7, the binding is strong, the fact that the novel coronavirus is highly infectious is met, and the experimental result is in line with the common knowledge.

(2) Sucrose is selected as the negative control of the experiment, and the concentration is 1 mg/mL; respectively preparing sample mixed solutions of spike protein (10000pg/mL) + sucrose (1mg/mL), spike protein (2000pg/mL) + sucrose (1mg/mL), spike protein (400pg/mL) + sucrose (1mg/mL), spike protein (80pg/mL) + sucrose (1mg/mL) and spike protein (16pg/mL) + sucrose (1mg/mL), putting the sample mixed solutions into sample bins 2-6 in sequence, putting a blank whole system buffer solution into bin 1, and putting a regenerated solution into bin 7; the Affinity program in BIAcore X100 was started and the assay was started for one cycle of Affinity.

As shown in FIG. 2, 5 mixed sample solutions with different concentrations were used to couple with the chip, and the binding parameters of the S protein and the ACE2 receptor in the negative control state were determined by fitting the coupling amounts of the S protein with different concentrations, and the analysis results are shown in Table 2.

TABLE 2

KD(M)	SE(KD)	Rmax(RU)	SE(Rmax)	offset(RU)	SE(offset)
						7.591E-7	3.9E-7
		898.8	5.8E+2	44.8	8.4

The binding constant KD (M) of the S protein and the ACE2 receptor in the negative control drug state is 7.591E-7, compared with the blank control 7.558E-7, the binding constant KD (M) is not significantly different, which indicates that sucrose cannot inhibit the binding of the S protein of the new coronavirus and ACE2, and the experimental result accords with the common general knowledge.

In the embodiment, the negative control adopts the lotus antipyretic capsule with the concentration of 1 mg/mL. Respectively preparing sample mixed solutions of spike protein (10000pg/mL) + lotus antipyretic capsule (1mg/mL), spike protein (2000pg/mL) + lotus antipyretic capsule (1mg/mL), spike protein (400pg/mL) + lotus antipyretic capsule (1mg/mL), spike protein (80pg/mL) + lotus antipyretic capsule (1mg/mL) and spike protein (16pg/mL) + lotus antipyretic capsule (1mg/mL), putting the sample mixed solutions into a sample bin 2-6, putting a blank system buffer solution into a bin 1, and putting a regenerated solution into a bin 7; the Affinity program in BIAcore X100 was started and the assay was started for one cycle of Affinity.

As shown in fig. 3, 5 mixed sample solutions with different concentrations were coupled to the chip, and the binding parameters of S protein and ACE2 receptor in the positive control state were determined by fitting the amounts of S protein coupled with different concentrations, and the analysis results are shown in table 3.

TABLE 3

KD(M)	SE(KD)	Rmax(RU)	SE(Rmax)	offset(RU)	SE(offset)
						1.300E-9	6.9E-10
		527.6	2.3E+2	-387.8	2.3E+2

The binding constant KD (M) of the S protein and the ACE2 receptor under the state of the positive control drug is 1.300E-9, and the situation that the lotus antipyretic capsule can block most of the ACE2 receptor binding region so that the butted S protein can easily reach balance, and the KD value is obviously reduced is understood.

In the experiment, nine drugs to be tested for the black pestilence-preventing medicine (the concentrated solution obtained in the preparation method, the crude drug content is 400g/L) are prepared into 1mg/mL aqueous solution, and then sample mixed solutions of spike protein (10000pg/mL) + nine black pestilence-preventing medicine aqueous solution (1mg/mL), spike protein (2000pg/mL) + nine black pestilence-preventing medicine aqueous solution (1mg/mL), spike protein (400pg/mL) + nine black pestilence-preventing medicine aqueous solution (1mg/mL), spike protein (80pg/mL) + nine black pestilence-preventing medicine aqueous solution (1mg/mL) and spike protein (16pg/mL) + nine black pestilence-preventing medicine aqueous solution (1mg/mL) are respectively prepared, sequentially placing the buffer solution into No. 1 bin and the regeneration solution into No. 7 bin; the Affinity program in BIAcore X100 was started and the assay was started for one cycle of Affinity.

As shown in fig. 4, 5 mixed sample solutions with different concentrations were coupled to the chip, and the coupling amount of S protein with different concentrations was used to measure the binding parameters of S protein and ACE2 receptor in the presence of the drug to be tested by fitting, and the analysis results are shown in table 4.

TABLE 4

KD(M)	SE(KD)	Rmax(RU)	SE(Rmax)	offset(RU)	SE(offset)
						7.327E-9	1.0E-9
		458.4	31	-102.7	34

The binding constant KD (M) of the S protein and an ACE2 receptor in the presence of the nine pestilence-preventing melanophores is 7.327E-9, and the nine pestilence-preventing melanophores can be understood to block most of the ACE2 receptor binding region, so that the docked S protein can easily reach equilibrium, and the KD value is obviously reduced compared with that of a blank control and a negative control.

TABLE 5 summary of the Activity

Name (R)	Kd(M)	SE(M)
			Blank control	7.558E-7	2.5E-7
Negative control (sucrose)	4.591E-7	3.9E-7
			Positive control (Lotus pestilence-clearing)	1.300E-9	6.9E-10
Nine-ingredient pestilence-preventing black medicine	7.327E-9	1.0E-9

The above are the binding parameters of the S protein and ACE2 under various conditions, and it can be seen from the above Table 5 that negative drug sucrose cannot affect the binding of ACE2 and S protein, and KD value is large and equivalent to that of blank control group; when the positive medicine lotus antipyretic capsule is added, the combination of ACE2 and S protein is sealed, and the KD value is reduced; the nine pestilence-preventing black medicines have better fusion inhibition activity on ACE2 target spots, and are closer to the positive medicine lotus antipyretic capsules.

ACE2, also known as achh, is known as angiotensin converting enzyme 2; the protein coded by the gene belongs to an angiotensin converting enzyme family of dipeptidyl carboxyl dipeptidase and has considerable homology with human angiotensin converting enzyme 1. ACE2 can regulate blood pressure, fluid balance, inflammation, cell proliferation, hypertrophy, and fibrosis; meanwhile, the organ and cell specific expression of the gene suggests that the gene may play a role in regulating cardiovascular and renal functions and in the aspect of birth. In this new corona epidemic situation, ACE2 allows the entry of new corona virus into cellular receptors, and studies have shown that small differences between the ACE2 receptor species can cause the inability of new corona virus to bind and infect the host. Therefore, ACE2 is the first key switch gene of host infected by new coronavirus, and if the receptor can be blocked, the new coronavirus can be blocked from infecting cells, so that the replication of the new coronavirus can be blocked; if the replication of the virus can be blocked, the reaction time can be won for the human immune system, and the new coronavirus infection can be prevented from expanding before the immune system is fully activated. The ACE2-SPIKE binding model is an important model for screening new crown drugs.

Second, complete skin irritation test

Test animals: common albino guinea pigs, female 3, weight 250- & ltSUB & gt 350g, order sheet number 20200029. Provided by the Dongxin Hua laboratory animal farm in the Guangzhou city Huadu district, the laboratory animal produces the license number: SCXK (Yue) 2019-0023; the skin was checked to be normal before the test.

The test conditions are as follows: license number for experimental animals: SYXK (Yue) 2019-.

The feed supplier cooperates with the creatures to produce the license number: suzuo Fed (2014) 01008. The temperature of the facility is 18-26 ℃, and the humidity is 40-70%.

The test method comprises the following steps: according to the skin irritation test 2.3.3 of the technical Specification for Disinfection (2002), hairs on two sides of the spine of the back of an animal are removed by a depilatory 24 hours before the test, so that the skin cannot be damaged; unhairing range, about 3cmx3cm left and right respectively; the next day, 0.5mL of the test substance was applied to one side of the skin and the other side was used as a blank control, and after 4h of application, the skin was washed with water or a suitable non-irritating solvent to remove the residue. Applying the composition once every day for 14d, observing the result 24h after each application, and scoring according to the scoring standard of skin irritation reaction shown in Table 2-9; in order to facilitate smearing and result observation of the test object, shearing is carried out if necessary; the control zone is treated in the same manner as the test zone.

And (3) detection results: the average score per animal per day was calculated based on the skin irritation response observation scores, and no irritation response occurred in the placebo zone, and the results of the test zone observations are shown in table 6.

TABLE 6 skin irritation response scoring results

As can be seen from Table 6, the average integral (irritation index) of each animal per day is 0, so that the skin irritation strength of the animals is evaluated according to the skin irritation strength grades in tables 2 to 9, and the nine-ingredient anti-plague hand sanitizer is nonirritating to the skin irritation strength of the animals for multiple times.

And thirdly, detecting the bacteriostasis of staphylococcus aureus and escherichia coli in the nine-ingredient plague-preventing hand sanitizer.

The detection basis is as follows: GB15979 supplement 2002 hygienic Standard for Disposable sanitary articles appendix C product bactericidal performance, bacteriostatic performance and stability test method; detection conditions are as follows: ambient temperature: 22.0 ℃, relative humidity: 50 percent.

And (3) detection results:

the results of 3 times of experiments show that the average bacteriostasis rate of the sample on staphylococcus aureus is more than 99.99 percent and the average bacteriostasis rate on escherichia coli is more than 99.99 percent after the sample acts for 2min (the results are shown in a table 7).

TABLE 7 bacteriostatic test results of nine-ingredient anti-pestilence hand-washing liquid

Note: negative control was grown aseptically.

And (4) conclusion: the nine-ingredient anti-plague hand sanitizer sample acts for 2min, has an average bacteriostasis rate of more than 99.99 percent on staphylococcus aureus and an average bacteriostasis rate of more than 99.99 percent on escherichia coli, and has strong bacteriostasis. Meets the regulation of GB15979-2002 sanitary standard of disposable sanitary products.

Fourth, bacteriostasis test of Candida albicans

Equipment: test strains: candida albicans ATCC 10231, the generation number of the above strain is 4, and a bacterial suspension is prepared by using PBS containing 0.03 mol/L.

The detection basis is as follows: GB15979 supplement 2002 hygienic Standard for Disposable sanitary articles appendix C product bactericidal performance, bacteriostatic performance and stability test method.

Detection conditions are as follows: ambient temperature: 22.0 ℃, relative humidity: 50 percent.

As a result: the results of 3 times of experiments show that the sample acts for 2min, the average bacteriostasis rate on the candida albicans is 60.40%, the sample acts for 5min, the average bacteriostasis rate on the candida albicans is 62.09%, the sample acts for 10min, the average bacteriostasis rate on the candida albicans is 64.24%, the sample acts for 20min, and the average bacteriostasis rate on the candida is 66.46% (see the results in table 8).

TABLE 8 bacteriostatic test results of nine-ingredient anti-pestilence hand-washing liquid

Note: negative control was grown aseptically.

Conclusion the sample, namely the nine-ingredient pestilence-preventing hand sanitizer, is used for 2min, has an average bacteriostasis rate of 60.40% on candida albicans, and has a bacteriostasis effect. Meets the regulation of GB15979-2002 sanitary standard of disposable sanitary products.

And fifthly, detecting mercury, arsenic and lead in the nine-ingredient plague-preventing hand sanitizer.

The detection basis is as follows: chapter iv physical and chemical inspection method of cosmetic safety technical Specification (2015 edition) 1.2 Mercury first method; 1.4 arsenic first method: 1.3 lead second method.

And (4) conclusion: the mercury content of the sample 'nine-ingredient anti-plague hand sanitizer' is less than 0.002mg/kg, the arsenic content is less than 0.11 mg/kg, and the lead content is less than 1.5 mg/kg.

Sixthly, stability test (two years biological method of validity period)

Test strains: staphylococcus aureus ATCC 6538, escherichia coli 8099. The generation number of the above strains is 4 th generation, and bacterial suspension is prepared by using PBS containing 0.03 mol/L.

The results of 3 times of experiments show that after the sample is stored for 90 days at 37 ℃, the sample acts for 2min, the average bacteriostasis rate on staphylococcus aureus is 98.83%, the average bacteriostasis rate on escherichia coli is 51.13%, the sample acts for 5min, the average bacteriostasis rate on staphylococcus aureus is 99.65%, the average bacteriostasis rate on escherichia coli is 57.15%, the sample acts for 10min, the average bacteriostasis rate on staphylococcus aureus is 99.92%, the average bacteriostasis rate on escherichia coli is 75.66%, the sample acts for 20min, the average bacteriostasis rate on staphylococcus aureus is more than 99.99%, and the average bacteriostasis rate on escherichia coli is 90.31% (the results are shown in table 9).

TABLE 9 bacteriostatic test results of nine-ingredient anti-pestilence hand-washing liquid

And (4) conclusion: after the sample of the nine-ingredient plague-preventing hand sanitizer is stored for 90 days at 37 ℃, the stock solution acts for 2min, the average bacteriostasis rate of the staphylococcus aureus is 98.83 percent, and the nine-ingredient plague-preventing hand sanitizer has strong bacteriostasis; the action lasts for 2min, the average bacteriostasis rate to the large intestine bacillus is 51.13 percent, and the antibacterial effect is achieved; the shelf life of the sample can be defined as 2 years according to GB15979-2002 hygienic Standard for Disposable sanitary articles appendix C6.

Claims (3)

1. The nine-ingredient plague-preventing hand sanitizer is characterized by being prepared from the following raw materials: 140-160 parts of fructus chebulae immaturus, 140-160 parts of aconitum pendulum, 140-160 parts of calamus, 140-160 parts of black sulfur, 140-160 parts of mukul myrrh, 140-160 parts of kyo ink, 4-6 parts of asafetida, 0.7-1.2 parts of musk and 65-75 parts of calculus bovis factitius.

2. The nine-ingredient pestilence-preventing hand sanitizer according to claim 1, wherein the hand sanitizer is prepared from the following raw materials: 150 parts of fructus chebulae immaturus, 150 parts of aconitum pendulum, 150 parts of acorus calamus, 150 parts of black sulfur, 150 parts of Mokul myrrh, 150 parts of Jingmo, 5 parts of asafetida, 1 part of musk and 70 parts of calculus bovis factitious.

3. The preparation method of the nine-ingredient plague-preventing hand sanitizer according to claim 1 or 2 is characterized by comprising the following steps: weighing fructus Chebulae Immaturus, radix Aconiti Szechenyiani, rhizoma Acori Calami, Sulfur, Myrrha, JINGJIU, resina Ferulae, Moschus, and artificial calculus bovis respectively to obtain mixed Chinese medicinal materials, and mixing the materials according to the ratio of medicinal materials-anhydrous ethanol: mixing the raw materials at a ratio of 1:30, refluxing for three times at 50 ℃, combining the supernatants of the three times, concentrating, drying and uniformly mixing to obtain a concentrated product with the crude drug content of 400-500 g/L, adding conventional auxiliary materials of the hand sanitizer into the concentrated product, and preparing the nine-ingredient anti-plague hand sanitizer according to a conventional method.

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