CN101843640A - Natural anti- virus and composition comprising thereof - Google Patents

Natural anti- virus and composition comprising thereof Download PDF

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Publication number
CN101843640A
CN101843640A CN201010183326A CN201010183326A CN101843640A CN 101843640 A CN101843640 A CN 101843640A CN 201010183326 A CN201010183326 A CN 201010183326A CN 201010183326 A CN201010183326 A CN 201010183326A CN 101843640 A CN101843640 A CN 101843640A
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virus
culture fluid
antiviral agent
weissella
lactobacillus
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CN101843640B (en
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李正华
张东逸
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COTDE Inc
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    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION 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
    • A01N63/00Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/20Bacteria; Substances produced thereby or obtained therefrom
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12Antivirals
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12Antivirals
    • A61P31/14Antivirals for RNA viruses
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12Antivirals
    • A61P31/14Antivirals for RNA viruses
    • A61P31/16Antivirals for RNA viruses for influenza or rhinoviruses

Abstract

Disclosed herein are a natural antiviral agent having high inhibitory activity against a broad spectrum of viruses without adverse side effects, obtainable using lactic acid bacteria isolated from kimchi, and a composition comprising the natural antiviral agent. The natural antiviral agent is safe to the body and can be applied to compositions, such as fodder, beverages, food, medicine, and cosmetics, imparting antimicrobial activity thereto.

Description

Natural anti-virus thing and the compositions that comprises it
The application is international application no PCT/KR2005/002348, international filing date on July 20th, 2005, China's application number 200580024612.3 enters Chinese January 22 2007 date, and denomination of invention is divided an application for " natural anti-virus thing and the compositions that comprises it ".
Technical field
The present invention relates to a kind of natural antiviral agent and the compositions that comprises it.More specifically, the present invention relates to the natural antiviral agent of a kind of application from the isolating lactobacillus preparation of Pickles (Kimchi), its height that shows at viral growth suppresses active, and be free from side effects, and to body is safe, the invention still further relates to compositions comprising described natural antiviral agent, such as beverage, feedstuff, food, medicine, cosmetics, or the like.
Background technology
Causing the influenza virus of influenza pandemic once every year, is a kind of alkaline single strand RNA virus, and its size with the described RNA of parcel is 1/10, the peplos of 000mm.According to antigenicity, influenza virus is divided into A, B and C type.Among them, influenza type A and B virus cause epidemic diseases.Be called hemagglutinin (HA) and neuraminidase (neuramidase) two types glycoprotein (NA), outstanding as projection from the surface, and 8 kinds of RNA fragments are present in virus inside.Two kinds of surface glycoprotein HA of influenza virus experience variation usually with NA in identical hypotype, this causes the appearance of new antigenic mutant in every year.
When cough, influenza virus sprays from pulmonary with the air of fierce release, and enters in other body by nose or mouth.In vivo, influenza virus is used viral glycoprotein h A projection, adheres to and penetrate the mucosal epithelium cell of upper respiratory tract, and here breeds subsequently.The result of study of accumulation shows viral infection cutter system really at present.Usually, on the target cell surface, virus and the receptors bind of forming by the sugar chain of glycoprotein, and experience endocytosis and the fusion of peplos and endosome subsequently, and enter target cell.At cell interior, virus by transcribing and duplicate them gene and produce progeny virion from their host's plasma membrane and breed.The infection of influenza virus be accompanied by usually fever, headache, arthralgia, whole body collapse, or the like, and have respiratory symptom, comprise sore throat, rhinorrhea, nasal obstruction, or the like.Described influenza is different from flu on the symptom and the order of severity, and is characterised in that most evil bad infectiousness, and this causes the fulminant epidemic diseases of short time in the whole world.Because influenza virus has the annual HA structure that all makes a variation, the antibody that produces so reply previous infection can not be effectively at occurent epidemic diseases, and this is a why popular reason of influenza.
For the infection of flu-prevention virus, scientists has planned and has been used to suppress to be attached to epithelial cell, penetrates cell, transcribes, duplicates, translates and discharges the method for filial generation from cell that all these is the target of resisiting influenza virus medicament.Under these methods, developed antiviral agent, such as amantadine, rimantadine, zanamivir etc., but, owing to it is reported that they produce significant adverse side effect in central nervous system, digestive system and autonomic nervous system, such as hypersensitivity, so must use extremely carefully.
As indicated above, the mucosal epithelium of influenza infection respiratory tract, and breeding there.If a kind of Virus Type of vaccine is different from described popular influenza virus, so inoculation may not can the infection and the propagation of flu-prevention virus effectively.In fact, not at effectively treating in the treatment of influenza virus or the prevention medicament.Now, often brushing teeth, keep throat moistening, and have a rest fully and nutrition, is to be known as the prevention method of effective influenza infection.
Owing to these reasons, exist for having the urgent need that height at influenza virus suppresses active and can not have the antiviral agent of adverse side effect ground application usually.
Virus disease is the danger of the biggest ground and destroys one of factor of poultry industry.Now, it is said that bird flu virus becomes the threat of the whole mankind and poultry industry.Recently, reported the outburst of H5N1 high pathogenic avian influenza (HPAI) in China and Southeast Asia birds continually.At brainstrust, the adaptation of bird flu virus height suddenlys change, rather than by radiation, this causes the fearful problem to the mankind from present situation.
In Indonesia, having been found that can not the measure of employing prevention property at the popular of birds influenza.In China, detected many H5N1 HPAI viruses, and they are to propagate than its control rate faster speed.Therefore, except inoculating, have no idea.In fact, inoculation is just carried out in many areas.
Vietnam and Thailand are in the more serious situation, have the birds influenza that increases number and have caused human dead report.Worsely, almost do not control the probability of influenza pandemic in these countries.Thailand, typical birds exporting country, the outburst of response bird flu is adhered to and will is in the policy that dangerous birds slaughters rather than inoculate.In order to eradicate bird flu, the Thai Government has been taked extreme measure, to without permission and immunity the personnel of poultry impose a fine.Reason is exactly, and can not eradicate bird flu in China because inoculation makes, Thailand has to stop the birds outlet.
In nearly all country with birds industry, fowl source infectious bronchitis (IB) is a representative disease of greatly destroying the home poultry raising industry.In practice, even some countries have eradicated newcastle (ND) or bird flu (AI), also there is not country to stop IB up till now.
In Korea S, the inspection data of the veterinary of country research and quarantine service organization (National Veterinary Researchand Quarantine Service) show, IB is most popular, and on the generation sum of poultry disease, it comprises 6% and comprised 5.2% in 2002 in calendar year 2001.IB virus is to study the most deep coronavirus, and has incredible simple structure.IB virus also is called removable virus, at present just in whole world experience sudden change.When they simultaneously during infection host, described sudden change is mainly caused by the gene recombinaton of serotype IB virus differing from each other.When the IB of two or more types virus while infection host, they are accompanied by the reorganization of their gene at identical Intracellular growth, and this causes having the appearance of the new virus of the feature that is different from those viral features of origin.For example, if gene recombinaton takes place between attenuated vaccine virus and pernicious wild-type virus, there is the probability that increases, promptly, the advantage of described two kinds of viruses, that is, and higher proliferation and pathogenic, with combination, and the progeny virus that is therefore obtained becomes the acute illness initiator of fast breeding most probably.Owing to this reason, vaccine, even attenuation, necessarily can not be introduced into domestic harmlessly.
The typical characteristics of IB virus comprises the lasting appearance of different serotypes and mutation, than the ability (18-36 hour incubation period) and the survival ability in birds colony (persistence in the farm) of the faster propagation of any other disease.In addition, defence and the cross immunity between the serotype at the serum antibody of described virus is limited.And, when two or more serotypes are attacked birds colony simultaneously, be difficult to only defend described virus with vaccine.
In Korea S, ND is caused by Avian pneumo-encephalitis virus, has been first kind of legal infectious disease by identification, and take place its every year in the hennery.Developed various vaccines, yet the generation of ND does not reduce still based on the ND inactivation of virus.
Now, the hennery has 3 defence lines that suppress bird disease.Article one, the defence line depends on bio-safety, and second defence line depends on inoculation, and the 3rd immune system of defending line to depend on birds self.Yet the generation of IB in the hennery proves that on the contrary IB virus crossed article one defence line of bio-safety, makes that inoculation is useless, and destroyed the health of chicken, destroyed immune system fully.Therefore, there is urgent need in the multifunctional treating agent for therapeutic agent that kills virus or unusual enhance immunity system.
Summary of the invention
Technical problem
Therefore, the present invention keeps it in mind the problems referred to above that exist in the prior art, and an object of the present invention is to provide novel natural antiviral agent, its height that has at various viral infection effectively suppresses active, and be the disadvantageous side effect that do not have to the human body tight security, described virus comprises influenza virus, bird flu virus, poultry infectious bronchitis virus etc.
In addition, another object of the present invention provides and contains described natural antiviral agent as composition of active components, and it can be contained in the various products.
Technical scheme
According to an aspect of of the present present invention, purpose of the present invention can realize like this, promptly, provide from being selected from by a kind of of the following group of forming the natural antiviral agent of preparation: the pickle fermentation process from the culture fluid of the isolating lactobacillus of Pickles, the concentrate of described culture fluid, the dehydrate of described culture fluid, and combination.
In described natural antiviral agent, lactobacillus can be selected from by Leuconostoc (Leuconostocspp.), Lactobacillus (Lactobacillus spp.), Wei Si Bordetella (Weissella spp.) and the group formed thereof.
Preferably, the Leuconostoc that is used for natural antiviral agent of the present invention comprises Fructus Citri Limoniae leukonid (Leuconostoc citreum), leuconostoc lactis (Leuconostoc lactis), Leuconostoc mesenteroides dextranicum (Leuconostoc mesenteroides subsp.Dextranicum), leuconostoc mesenteroides subsp mesenteroides (Leuconostoc mesenteroides subsp.Mesenteroides), Argentina leukonid (Leuconostoc argentinum), yellowish pink leukonid (Leuconostoc carnosum), hard leukonid (Leuconostoc gellidum), Leuconostoc kimchii (Leuconostoc kimchii), Leuconostoc inhae, Leuconostoc gasicomitatum, and their combination.
Preferably, the Lactobacillus that is used for natural antiviral agent of the present invention comprises Lactobacillus brevis (Lactobacillus brevis), bacillus acidophilus (Lactobacillus acidophilus), Lactobacillus bulgaricus (Lactobacillus bulgaricus), Lactobacillus plantarum (Lactobacillusplantarum), Pickles lactobacillus (Lactobacillus kimchii), class Lactobacillus plantarum (Lactobacillus para-plantarum), lactobacillus curvatus subspecies (Lactobacillus curvatussubsp.Curvatus), Lactobacillus sakei subsp.Sakei, and their combination.
Preferably, the Wei Si Bordetella that is used for natural antiviral agent of the present invention comprise Weissellakoreens, Weissella hanii, Pickles Wei Si Salmonella (Weissella kimchii), Weissella soli, merge Wei Si Salmonella (Weissella confusa), and their combination.
According to another aspect of the present invention, purpose of the present invention can realize as composition of active components by a kind of natural antiviral agent that comprises is provided, described natural antiviral agent prepares from being selected from by a kind of of the following group of forming: the pickle fermentation process from the culture fluid of the isolating lactobacillus of Pickles, the concentrate of described culture fluid, the dehydrate of described culture fluid, and combination.
In this, the described compositions form of one of beverage, feedstuff, food, medicine or cosmetics preferably.
Advantageous effects
The culture fluid of pickle lactic acid bacteria, or their concentrate or dry matter have the fabulous antimicrobial acivity at wide spectrum virus, and can give food, feedstuff, beverage, medicine and cosmetics safety effective antiviral activity.
Implement optimal mode of the present invention
About the present invention, the inventor carry out about foods and cosmetics preserve deeply and thorough research causes such discovery, promptly, can act as multi-functional natural antiviral agent from the isolating lactobacillus of Pickles (pickle lactic acid bacteria hereinafter referred to as) culture, it has at the effective inhibition of wide spectrum virus active.Known ground, pickle lactic acid bacteria is bred in the pickle fermentation process, it is usually by using the pickled Rhaphanus of salt, Brassica oleracea L.var.capitata L. and Fructus Cucumidis sativi, described souse is mixed with the flavoring compositions that comprises Fructus Capsici powder, mashed garlic, green Bulbus Allii Cepae end, the briny seafood of salt etc., and described pickless fermented prepare.
The invention pattern
Better understanding of the present invention can obtain by following example, and described example proposes for illustrating, but can not be interpreted as restriction of the present invention.
Embodiment
Preparation pickle lactic acid bacteria culture fluid
In pickle lactic acid bacteria, Leuconostoc kimchii (KCTC2386), leuconostoc mesenteroides subsp mesenteroides (KCTC3530), Leuconostoc citreum (KCTC3524), leuconostoc lactis (KCTC3528), Lactobacillus plantarum (KCCM11322) and Weissella koreensis (KCTC3746) are used to prepare described culture fluid.
At first, 100ml MRS (Difco) meat soup 121 ℃ of autoclavings 15 minutes, and is inoculated into pickle lactic acid bacteria in the meat soup, and 30 ℃ of wave and culture 16 hours.
, behind the centrifugal 3min of 5000 * g, the supernatant is filtered by the membrane filter with 0.22 oral pore size at the culture that will be obtained, to obtain sterile solution.
Subsequently, by having 10,000Da holds back the film of size with this sterile solution.With having 1,000Da holds back the membrance concentration of size with resulting filtrate.The concentrate that obtains is like this filtered by the membrane filter with 0.22 oral pore size again, so that keep aseptic condition.In order to get rid of the pH-dependency effect of antimicrobial acivity, final culture fluid is adjusted to pH 7.
-detect the pickle lactic acid bacteria culture fluid at the antiviral activity of bird flu virus-
The animal cell line (MDCK (Madin-Darby dog kidney) or CEF (Embryo Gallus domesticus fibroblast)) of monolayer growth is infected with bird flu virus HK/220, cultivated then 3 days.Plaque forms has proved viral infection.
To be diluted to 10 with PBS (pH 7.4) -3Bird flu virus and with the mixture of the pickle lactic acid bacteria culture fluid of identical buffer salt dilution, add on flat board with monolayer growth converged 80% or more animal cell line MDCK or CEF in, subsequently described cell line was cultivated 3 days.With the cell fixation of infection and with dimethyl diaminophenazine chloride dyeing, to observe the wherein formation of plaque.The result provides in table 1, and is as follows.
Table 1
Figure GSA00000124139200071
Apparent from the data of table 1, do not allow to reach 1.3~1.6 * 10 although do not contain the contrast of pickle lactic acid bacteria culture fluid 2Individual plaque forms, and has reduced the plaque number that forms significantly but add Pickles dish lactic acid bacteria culture solution.Therefore, these results hint that described Pickles dish lactic acid bacteria culture solution has very effective antiviral activity at bird flu virus.
-detect the pickle lactic acid bacteria culture fluid at the antiviral activity of influenza virus-
Replace the bird flu virus HK/220 except using influenza virus A 1H3N2, repeat and identical step in the antiviral activity at bird flu virus detects.Described result provides in table 2, and is as follows.
Table 2
Figure GSA00000124139200072
What do not have described pickle lactic acid bacteria culture fluid although wherein only used the viral dilution thing has 1.3~1.6 * 10 to impinging upon each culture plate 2Individual plaque, as shown in table 2, but when having the pickle lactic acid bacteria culture fluid, the infection of influenza virus has developed the plaque of remarkable minimizing number, and this hints that described pickle lactic acid bacteria culture fluid has very effective antiviral activity at influenza virus.
The animal detection of-pickle lactic acid bacteria culture fluid-
1,100 hen of laying eggs of the egg production of minimizing is divided into 10 groups at random to take place experience significantly in the time of will be owing to bird flu and newcastle, and every group comprises 110 hens.The pickle lactic acid bacteria culture fluid of above preparation is diluted 10 times in the hen drinking water, and in 17 days process, observe the variation of egg production.The result provides in following table 3.
In the table 3, sample 1 prepares based on Leuconostoc kimchii (KCTC2386) culture fluid, sample 2 is based on leuconostoc mesenteroides subsp mesenteroides (KCTC3530) culture fluid, sample 3 is based on Leuconostoccitreum (KCTC3524) culture fluid, sample 4 is based on leuconostoc lactis (KCTC3528) culture fluid, sample 5 is based on Lactobacillus plantarum (KCCM11322) culture fluid, sample 6 is based on Weissellakoreensis (KCTC3746) culture fluid, sample 7 is based on the mixture of leuconostoc mesenteroides subsp mesenteroides (KCTC3530) culture fluid and Lactobacillus plantarum (KCCM11322) culture fluid, sample 8 is based on the mixture of Leuconostoc kimchii (KCTC2386) culture fluid and Weissella koreensis (KCTC3746) culture fluid, and sample 9 is based on the mixture of Weissella koreensis (KCTC3746) culture fluid and Lactobacillus plantarum (KCCM11322) culture fluid and prepare.
Table 3
Figure GSA00000124139200081
As shown in table 3, reduce owing to infective virus causes egg production, suffer from dysentery and show the bird inlay of unusual respiratory symptom, drink to them contain according to the water of culture fluid of the present invention after 10 days, return to normal condition, this determines by the level that egg production returns to normal hen.Therefore, also has fabulous interior resisting virus activity according to pickle lactic acid bacteria culture fluid of the present invention.
In addition, owing to natural antiviral agent of the present invention is to prepare from eugonic pickle lactic acid bacteria culture fluid between the food Pickles yeast phase of Korean traditional, therefore, detect, there is not suspection in the safety of body about described natural anti-virus product even without carrying out extra safety.
As in the drinking water of bird inlay, even without carrying out concrete detection, the culture fluid of antiviral activity of the present invention also can be included in food, feedstuff, beverage, cosmetics and the medicine.
When culture fluid of the present invention is used for cosmetics, beverage, food, feedstuff, medicine etc., it should be appreciated by those skilled in the art, according to the characteristic of described product and the embodiment of preparation, distribution, storage and the application of described product, various modifications, interpolation and replacement be can in described culture fluid, carry out, and the present invention scope of disclosure and spirit in the claim of enclosing do not deviated from.
Industrial usability
In sum, the data that obtain in an embodiment show that the pickle lactic acid bacteria nutrient solution has the fabulous antiviral activity for broad-spectrum micro-organisms, and can give food, feed, beverage, medicine and cosmetics safety and effective antiviral activity.

Claims (2)

1. compositions, it comprises from the natural antiviral agent that is selected from by a kind of of the following group of forming preparation as active component: the pickle fermentation process from the isolating Wei Si Bordetella of Pickles (Weissella spp.) culture fluid, the concentrate of described culture fluid, the dehydrate of described culture fluid, and their combination.
2. the compositions that proposes of claim 1, wherein said Wei Si Bordetella comprise Weissellakoreensi, Weissella hanii, Pickles Wei Si Salmonella (Weissella kimchii), Weissella soli, merge Wei Si Salmonella (Weissella confusa), and their combination.
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