Technical background:
Bird flu is a kind of acute infectious disease that is caused by avian influenza virus, also can infect the mankind, metainfective symptom mainly shows as high heat, cough, runny nose, myalgia etc., and is most with serious pneumonia, multiple organ failures such as the severe patient heart, kidney cause death, and case fatality rate is very high.This disease can be passed through number of ways propagation such as digestive tract, respiratory tract, and personnel and vehicle contact are to propagate this sick key factor.
Some national H5N1 avian flu casess are obvious expansion situation, and situation is very complicated.International organizations such as The World Health Organization (WHO) give a warning a few days ago, and bird flu epidemic situation has the danger that breaks out on a large scale, call upon States to enhance your vigilance, and prepare to respond.Migrate season migratory bird, the whole world has three migratory birds to migrate route through China, and the band poison is migrated migratory bird and contacted with poultry, causes new epidemic situation easily.China's some areas poultry rearing mode is backward relatively, and live-bird circulation and trans-regional allocation and transportation are frequent, and also there are some weak links in prevention and control.China's highly pathogenic bird flu prevention and control task is quite arduous.
The bird flu in Hong Kong in 1997 has caused the no small influenza of scale one time, in current influenza pandemic process 6 people's death is arranged.The Hongkong government decides on the spot and has destroyed the whole poultry that comprise chicken.Current Mao flu and the flu outbreak that took place in 1918 were separated by nearly 80 years, study through scientist, although twice influenza pandemic is remote at interval, but similarity is arranged: the genetic mutation that influenza virus is taken place a kind of human immune system that recombinates out does not have contacted influenza virus new variant, and influenza is transferred on the person on one's body from animal then.
To the treatment of bird flu, still lack desirable specific medicament at present.In the human avian influenza diagnosis and treatment scheme that ministry of Health of China is printed and distributed, main Oseltamivir (the Oseltamivir that adopts, Tamiflu), amantadine (Amantadine) and Rimantadine antiviral drugs such as (Rimantadine), be aided with symptomatic treatment and Chinese herb decoction treatments such as analgesic expelling phlegm and arresting coughing.
We adopt pure Chinese medical extract: the extract of oil of zedoary turmeric and hypericum perforatum is formed compound preparation, can provide method preferably for the prevention and the treatment of bird flu.Contain rcumenol in the oil of zedoary turmeric, (RSV) has direct repression to Respiratory Syncytial Virus(RSV), and influenza virus A1 type and A3 type are had direct deactivation.Hypericum perforatum is the herb or the whole herb with root of dicotyledon medicine Guttiferae plant hypericum perforatum, hypericin that wherein contains (Hypericin) and pseudohypericin (pesudohypericin) have very strong antivirus action, in in vitro test, can suppress miscellaneous retroviruses (retrolirus) consumingly.These two kinds of drug combinations can improve curative effect, and safety simultaneously is also better.
Summary of the invention:
The present invention aims to provide a kind of antivirus sprayer and preparation method thereof.This spray both can suppress or kill pathogenic microorganisms such as body surface, article and airborne virus, bacterium, can spray into prevention and treatment viral infection disease through the bird mouth again.
Concrete technical scheme is as follows:
By the compound spray that oil of zedoary turmeric and Hypericum Perforatum P.E are formed, this prescription has significant antiviral antibacterial action, and its concrete feature is as follows:
The oleopene that the zingiberaceous plant curcuma zedoary extracts is made active component one oil of zedoary turmeric, and the wounded in the battle herb extract of Guttiferae plant hypericum perforatum HypericumPerforatum is mixed with solid or liquid or sterile pharmaceutical formulation according to a certain percentage.
1, this compound antivirus sprayer is characterized in that, prescription (weight portion) is:
Oil of zedoary turmeric 0.1~10%
Hypericum perforatum shifts to an earlier date thing 0.1~10%
Cosolvent A 1~5%
Cosolvent B 1~5%
Bacteriostatic agent 0.01~0.5%
Distilled water adds to 100%
2, as claims 1 said spray, it is characterized in that described cosolvent A and B are selected from: fatty acid sorb smooth (SPan) class material, polysorbate (Tween) class material, polyoxyethylene fatty acid ester class material, polyoxyethylene aliphatic alcohol ether (Brij) class material, polyoxyethylene one polyoxypropylene copolymer, lauryl sodium sulfate or cetyltrimethylammonium base amine bromide or its mixture.
3, as claims 1 said spray, it is characterized in that described bacteriostatic agent is selected from: parabens (methyl esters, propyl ester), benzalkonium bromide, phemerol chloride, phenmethylol, anesin, thimerosal etc.
4, a kind of preparation method of compound antivirus sprayer is characterized in that, is prepared according to the following steps:
(1) takes by weighing the oil of zedoary turmeric of the described amount of claim 1, add cosolvent A and mix, be solution 1;
(2) hypericum perforatum that takes by weighing the described amount of claim 1 shifts to an earlier date thing, adds cosolvent B and mixes, and is solution 2;
(3) solution 2 under agitation is added in the solution 1, and mixes;
(4) bacteriostatic agent behind a small amount of dissolved in distilled water is joined in the above-mentioned mixed liquor, stir;
(5) adding distil water carries out intermediate and detects to about full dose, and adjusting pH value and content are qualified to product;
(6) soup elder generation filter coarse filtration under 10,000 grades of purification conditions with the smart filter of ultrafilter, is divided in the container that has sprayer more then;
(7) check: the finished product after the packing is carried out the examination and test of products, then warehouse-in.
Good effect of the present invention: this pharmaceutical preparation can form with spraying in micro-sprayer is used.(1) is used for the surrounding air sterilization, can effectively kill various viruses, germ in the short time, dye, do not have corrosion to human body is nontoxic, harmless, colourless, do not need rinsing after the spraying, also can not produce drug resistance, the using method uniqueness portably uses very simple, can be people clean live and work environment is provided whenever and wherever possible, away from the infringement of virus and germ; (2) can spray into prevention and treatment viral infection disease through the bird mouth again.
Embodiment 1:
Prescription:
Oil of zedoary turmeric 0.3%
Hypericum perforatum shifts to an earlier date thing 0.25%
Tween 80 2.5%
Ethanol 2.5%
Propylben 0.02%
Distilled water adds to 100%
Compound method:
(1) takes by weighing the recipe quantity oil of zedoary turmeric, add Tween 80 and mix, be solution 1;
(2) hypericum perforatum of the place's of taking by weighing amount shifts to an earlier date thing, adds the recipe quantity dissolve with ethanol, is solution 2;
(3) solution 2 under agitation is added in the solution 1, and mixes;
(4) propylben behind a small amount of dissolved in distilled water is joined in the above-mentioned mixed liquor, stir;
(5) adding distil water carries out intermediate and detects to 95% of about full dose, and adjusting pH value and content are qualified to product;
(6) soup elder generation filter coarse filtration under 10,000 grades of purification conditions with the smart filter of ultrafilter, is divided in the container that has sprayer more then;
(7) check: the finished product after the packing is carried out the examination and test of products, then warehouse-in.
Embodiment 2:
Prescription:
Oil of zedoary turmeric 1%
Hypericum perforatum shifts to an earlier date thing 0.5%
Ethanol 5%
Thimerosal 0.01%
Distilled water adds to 100%
Compound method:
(1) hypericum perforatum of the place of taking by weighing amount shifts to an earlier date thing, adds half dissolve with ethanol of recipe quantity, is solution 1;
(2) take by weighing the recipe quantity oil of zedoary turmeric, add half ethanol of recipe quantity and stir, be solution 2;
(3) solution 2 under agitation is added in the solution 1, and mixes;
(4) thimerosal behind a small amount of dissolved in distilled water is joined in the above-mentioned mixed liquor, stir;
(5) adding distil water carries out intermediate and detects to 95% of about full dose, and adjusting pH value and content are qualified to product;
(6) soup elder generation filter coarse filtration under 10,000 grades of purification conditions with the smart filter of ultrafilter, is divided in the container that has sprayer more then;
(7) check: the finished product after the packing is carried out the examination and test of products, then warehouse-in.
The experiment of anti-chicken source H5-AIV virus:
1 materials and methods
1.1 material
1.1.1 test virus of A IV Strain: A/Chicken/Guangdong/1/2004 (being called for short chicken source H5-AIV) is by the separation of veterinary college poultry diease teaching and research room of Agricultural University Of South China, purification storage.
1.1.2 test is 20 μ g/mL, 200 μ g/mL, 500 μ g/mL with the concentration of medicine and source Hypericum Perforatum P.E solution; The concentration of oil of zedoary turmeric solution is 20 μ g/mL, 200 μ g/mL, 500 μ g/mL; 1. sample is according to embodiment 1 prescription and technology preparation; 2. sample is according to embodiment 2 prescriptions and technology preparation.
1.1.3 test is a low immune chicken embryo in 10 day age with the chicken embryo, Co., Ltd provides by the yellow herding in hilllock, ruins, Foshan City;
1.1.4 HA test is 0.5% chicken red blood cell with red blood cell, prepares voluntarily;
1.1.5 test soda acid 0.1N NaOH, 0.1N HCl.
1.2 method
1.2.1 the preparation of 0.5% chicken red blood cell
By wing vein or the heart blood sampling of the cock that grows up, put into sterilization test tube (add 3.8% sterilization sodium citrate 0.2mL by every milliliter of blood and make anticoagulant), rapidly mixing.Then blood is injected centrifuge tube, through 3000 rev/mins, centrifugal 10~15min, inhale the leucocyte film that goes to supernatant and red blood cell upper strata with suction pipe, the red blood cell that precipitates is added dilution (physiological saline) washing, again with the centrifugal 10~15min abandoning supernatant of centrifuge 3000 commentaries on classics/min, add the dilution washing again, so cyclic washing is three times, with last after centrifugal red blood cell mud, the dilution factor adding dilution (the 0.5mL red blood cell adds 99.5mL physiological saline) by 0.5%.
1.2.2 avian influenza virus chicken embryo median infective dose (EID
50) mensuration
Virus stock solution used is pressed 10 with sterile saline
-3~10
-1010 days instar chicken embryos are inoculated in dilution, each dilution factor inoculation 8 pieces of embryo (chicken source H5-AIV has adopted the method for 7 pieces of embryos of each dilution factor), and the 0.2mL/ embryo is hatched for 37 ℃, and chicken embryo dead in 24 hours discards.Measure that the HA of dead embryo tires after 24 hours, positive greater than 4 holes.Press Reed-Muench Liang Shi method and calculate EID
50
1.2.3 the adjustment of medicine pH value
Use 0.1N NaOH, 0.1N HCl is adjusted to the pH value of anti-avian influenza virus Chinese medicine between 6~8.
1.2.4 medicine is to the mensuration of the half toxic dose of chicken embryo
The doubling dilution medicine is established 5 degree, 5 embryo/groups, and the 0.2mL/ embryo, 96 hours, other established the blank group.Embryo is not caused death, do not influence idiosome normal development person and be judged to safety.Press Reed-Muench Liang Shi method and calculate the half toxic dose, as using the equal normal development of medicine stoste group idiosome, visual original liquid concentration is the half toxic dose.
1.2.5 medicine is to the inhibitory action of avian influenza virus
Inoculate 50 EID
50The medicine of virus (0.2mL)+maximum safe dose (0.2mL), 5 pieces/group; 37 ℃, 72 hours, embryo dead in 24 hours discarded.72h freezes embryo (dead embryo then freezes embryo immediately behind the 24h), surveys HA and tires, and is positive greater than 4 holes.Other establishes the virus-positive control group.
2 results and analysis
2.1 avian influenza virus chicken embryo median infective dose (EID
50) measurement result such as table 3.1
Table 3.1 avian influenza virus chicken embryo median infective dose (EID
50)
Virus liquid dilution factor |
Chicken source H5-AIV |
10
-3 10
-4 10
-5 10
-6 10
-7 10
-8 10
-9 10
-10 EID
50 |
7/7
* 7/7 7/7 7/7 3/7 1/7 0/7 0/7 10
7.338/0.2mL
|
*Be the positive embryo number of HA/inoculation embryo number
2.2 medicine is to the measurement result such as the table 3.2 of the half toxic dose of chicken embryo
The mensuration of table 3.2 half toxic dose
Experimental group |
Drug concentration |
Half toxic dose (mg) |
Maximum safe dose (μ g/mL) |
20μg/m L |
200μg/m L |
500μg/m L |
250μg/mL +0.3% |
500μg/mL +1% |
1. sample is 2. for hypericum perforatum oil of zedoary turmeric sample |
5/5
* 4/5 / /
|
5/5 5/5 / / |
5/5 5/5 / / |
/ / 5/5 / |
/ / / 5/5 |
500 500 250μg/mL+0.3 % 500μg/mL+1% |
500 500 250μg/mL+0.3 % 500μg/mL+1% |
*Be normal development embryo number/connect embryo sum
2.3 medicine is to the inhibitory action result of avian influenza virus
2.3.1 medicine is to inhibitory action result such as the table 3.3 of chicken source H5-AIV
Table 3.3 medicine is to the inhibitory action result of chicken source H5-AIV
Experimental group |
HA (the Log that tires
2)
|
Infect ratio |
Inhibiting rate (%) |
Hypericum perforatum oil of zedoary turmeric sample is 2. positive control of sample 1. |
0 6 0 0 10 |
0 0 0 0 8 |
0 0 0 0 10 |
0 8 0 0 9 |
0 0 0 0 10 |
0 5 0 0 10 |
0 0 0 0 10 |
5 0 0 0 8 |
0 9 0 0 8 |
5 0 0 0 10 |
2/10=0.2 4/10=0.4 0/10=0 0/10=0 10/10=1 |
80 40 100 100 / |
2.3.5 all test Chinese medicines and compounds are to the inhibiting result of the chicken source H5-AIV strain result according to table 3-3, tire with the equal HA of allantoic fluid in 10 pieces of embryos of "+" expression and to be 0, tire all greater than 4 holes with the equal HA of allantoic fluid in 10 pieces of embryos of "-" expression expression, tiring with the allantoic fluid HA of part embryo in 10 pieces of embryos of " ± " expression is 0, and "/" expression is not used as corresponding test.Table 3.4 is the inhibiting result of all test Chinese herbal medicines to chicken source H5-AIV strain.
All test Chinese medicines of table 3.4 and compound are to the inhibiting result of chicken source H5-AIV strain
Experimental group |
Chicken source H5-AIV |
1. sample is 2. for hypericum perforatum oil of zedoary turmeric sample |
± ± + + |