The composite bacteriostatic agent of a kind of plant polyphenol
Technical field
The invention belongs to the Prevention Technique field of plant pest.Particularly, the present invention relates to a kind of plant polyphenol bacteriostatic agent and preparation method thereof.
Background technology
Be widely used in the bactericide on trees and the plant at present, require to have higher bactericidal properties and persistence preferably, protect trees and plant is not subjected to the infringement of bacterium worm to reach.
This series bactericidal agent is still based on chemical pesticide.But bacterium worm fertility is strong, and is fast to environmental adaptation, Multiple Pesticides produced pesticide resistance.In addition, unreasonable use chemical pesticide, uses or excess is used chemical pesticide continuously for example single use, can quicken speed and development degree that the bacterium worm develops immunity to drugs again.Along with the drug-fast enhancing of pest, people constantly strengthen the agricultural chemicals using dosage aborning, so formed the strong dependence to agricultural chemicals.Although it is important that some chemicalses have play a part aspect bactericidal, guaranteed the stable and high yields of agricultural products, the high toxicity of these chemicalses, serious residual and pollution problem have also brought great hidden danger to human health.Especially today, people more and more pay attention to environmental problem, more and more need more pollution-free food, therefore, solve high toxicity, high residue is very urgent with pollution problem.For reaching this purpose, that people pay attention to seeking from animals and plants and microorganism more is efficient, the new type bactericide of low toxicity, low-residual or noresidue.
Plant polyphenol is that a class extensively is present in the polyphenols in the plant corpus, and the content in plant is only second to cellulose, hemicellulose and lignin, mainly is present in the skin, root, wood, leaf, fruit of plant.Its complex structure; chemical property is active, and often exists with the form of mixtures of the homologue of a large amount of similar performances, and plant polyphenol is not only resisted the infringement of herbivores to plant corpus; simultaneously also stoped infecting of microorganism, comprised tolerance and the protection injury of raising various pathogenic bacteria.Except plant pathogen, plant polyphenol in the animal body and other environment in microorganism can both produce the obvious suppression effect, the microorganism that comprises fungi, bacterium, yeast and virus etc. is had resistance of wide spectrum.
The ability of plant polyphenol height conjugated protein is undoubtedly a principal element of its biocidal property, tannin often just shows the obvious suppression effect at low concentration very, it is to pass through inhibitory enzyme activity, destroy the pathogen metabolic process and realize that it is inhibiting, in addition, it can also by combine with cell membrane or with the microorganism living environment in complexing of metal ion, destroy the eubolism of pathogen, thereby also can realize its inhibitory action.The chemical constitution of plant polyphenol and molecular weight influence its activity.The number of phenolic hydroxyl group on phenyl ring and the relative toxicity of position influence aldehydes matter, the degree of hydroxylation enhancing causes toxicity to strengthen.The big person of molecular weight does not often demonstrate biocidal property, and low-molecular-weight polyphenol often shows stronger microorganism toxicity.
But along with prolonged and repeated application, germ has produced resistance in various degree gradually to medicament.In order to overcome or the generation of delaying drug resistance, prolong medicament service life, enlarge antibiotic enhancing and press down the effect of killing germ, the fungicide compounding of the different modes of action, the different mechanisms of action is used, be one of effective measures that solve the delaying drug resistance development.
In order to overcome these problems that exist in the prior art, the inventor is according to the characteristics of plant polyphenol, carried out a large amount of research and test, it is composite to utilize plant polyphenol and Radix Glycyrrhizae to carry out, thereby develops the composite bacteriostatic agent of the natural environmental-protective type that can overcome the chemical pesticide defective.
Summary of the invention
Problem to be solved
The purpose of this invention is to provide a kind of composite bacteriostatic agent of plant polyphenol that overcomes above-mentioned chemical bactericide defective.
Another object of the present invention is the preparation method of the composite bacteriostatic agent of a kind of above-mentioned plant polyphenol.
Technical scheme
Valonea extract be by the shell rubber Bowl of wild plant through leaching, vacuum evaporation concentrate, atomized drying and the mixture of a kind of natural poly-phenol organic compound of making.Main component is a hydrolysable tannins, and its structure is very complicated, is to have phenol formula structure, quinoid structure and polyhydric compound.The physical property of this product is a bitter and puckery flavor, and convergence is arranged, and outward appearance is amorphous chocolate brown powder.Chemical property is very active.Hydrolysable tannin major part wherein is an Ellagitannins, belongs to C glycosidic bond connected mode, the big and complex structure of its molecular weight, and hydrolysis and ester exchange are all incomplete.Can't adopt normally used hydrolysis to make the tannin cracking of molecules, reach the purpose that reduces its molecular weight.
Radix Glycyrrhizae (Glycyrrhiza uralensis), leguminous plant, perennial herbaceous plant is China's traditional Chinese medicine, has detoxifcation, anti-inflammatory, effect such as antitumor grade, wherein medicinal active ingredient is glycyrrhizic acid, enoxolone and multiple flavone component.Flavonoid substances is another kind of important physical active substance in the Radix Glycyrrhizae, at the Radix Glycyrrhizae antiulcer, separate in the physiological functions such as spasm and played important function.Licoflavone class material has comprised glycyrrhizin, liquiritin, isoliquiritin, Neoliquiritin, Neoisoliquiritin, isoliquiritigenin-4-β-glucose-β-appropriate glucosides of foreign Yuan etc.Also contain the licorice polysaccharide than horn of plenty in the Radix Glycyrrhizae, it is a kind of antiviral ingredients in the Radix Glycyrrhizae.Similarly natural products can also be mentioned ginkgo leaf powder, scutelloside phenol etc.For for simplicity, on meaning of the present invention, Radix Glycyrrhizae, ginkgo leaf powder, scutelloside phenol are referred to as natural products.
Composite bacteriostatic agent of plant polyphenol of the present invention and preparation method thereof will be described in detail belows.
The composite bacteriostatic agent of a kind of plant polyphenol, it is characterized in that it by one or more be selected from the tannin extract of valonea extract, larch extract, Acacia extract and one or more natural productss that is selected from licorice powder, ginkgo leaf powder, scutelloside phenol with dry weight basis according to weight ratio 0.5-1.5: 1.5-2.5 forms.
Preferably, the composite bacteriostatic agent of plant polyphenol of the present invention by one or more be selected from the tannin extract of valonea extract, larch extract, Acacia extract and one or more natural productss that is selected from licorice powder, ginkgo leaf powder, scutelloside phenol with dry weight basis according to weight ratio 0.8-1.2: 1.8-2.2 forms.
More preferably, the composite bacteriostatic agent of plant polyphenol of the present invention is selected from by one or more that tannin extract of valonea extract, larch extract, Acacia extract and one or more natural productss that is selected from licorice powder, ginkgo leaf powder, scutelloside phenol form according to weight ratio with dry weight basis at 1: 2.
In addition, the present invention relates to the composite bacteriostatic agent of a kind of plant polyphenol, it is characterized in that it is made up of with licorice powder and/or ginkgo leaf powder valonea extract and/or larch extract.
Preferably, the composite bacteriostatic agent of plant polyphenol of the present invention is characterized in that it is made up of valonea extract and licorice powder.
In addition, the present invention relates to the preparation method of the composite bacteriostatic agent of a kind of plant polyphenol, it is characterized in that this method is that one or more are selected from the tannin extract of valonea extract, larch extract, Acacia extract and the natural products aqueous solution that is prepared into soluble in water that one or more are selected from licorice powder, ginkgo leaf powder, scutelloside phenol, this aqueous solution contains the described tannin extract in this aqueous solution gross weight 1.5-2.5%, the described natural products of 3.0-5.0%.
Described tannin extract can buy from each tannin extract factory; Described natural products can directly buy from biopharmaceutical company, and purity is more than 80%.Described tannin extract and natural products are pulverous, restriction that its granularity has nothing special.
Described dissolving can be carried out under room temperature and normal pressure, also can suitably heat during dissolving, but temperature generally is no more than 50 ℃.Described water can be running water, distilled water, river, well water etc.Described tannin extract and natural products have no particular limits in the order that dissolving adds in the entry time, promptly can be earlier with tannin extract water-soluble after, water-soluble natural products again, perhaps vice versa, perhaps adds in the entry simultaneously and dissolve.After dissolving fully, described tannin extract and natural products promptly obtain the composite bacteriostatic agent of plant polyphenol of the present invention.
Preferably, this aqueous solution contains the described tannin extract in this aqueous solution gross weight 1.8-2.2%, the described natural products of 3.5-4.5%.
More preferably, this aqueous solution contains in this aqueous solution gross weight 2.0% described tannin extract, 4.0% described natural products.
Beneficial effect
According to the characteristics of the chemical constitution of plant polyphenol, develop bacteriostatic agent of the present invention, not only have antibacterial effect, also have the effect of protective plant.
Since plant polyphenol utilize especially plant polyphenol in the animal body and other environment in microorganism can both produce the obvious suppression effect; the microorganism that comprises fungi, bacterium, yeast and virus etc. had resistance of wide spectrum; it is not a kill pathogens; but the survival of inhibition germ; reach the effect of protective plant, this is the unique aspect of this bacteriostatic agent.The present invention directly selects each tannin extracts for use, directly and other natural products carry out composite.
Secondly, polyphenol has the effect of heatstroke prevention resisting cold, can reduce ultraviolet irradiation, and protective plant is not subjected to the infringement of photo-thermal; Can help the function of branches and leaves defence frost in addition, so have the effect of protective plant.
Plant polyphenol bacteriostatic agent of the present invention can effectively prevent general fungi on the plant and the infringement of bacterium.As: the bacterium that can cause dothiorella gregaria; Can cause chestnut epidemic disease, claim the bacterium of die back disease, canker, hole rot again.Colonize on the plants such as chestnut class, oak class, lacquer tree, fagus, main harm corn, rice etc. cause ear rot and make rice mouldy, cause the bacterium of pestalotiopsis funerea; The main harm coniferous species is with the bacterium of masson pine, wet-land pine tree, pinus yunnanensis, torch pine, Chinese pine etc.
Have high-drug-effect, low toxicity, low-residual, nuisanceless; Have paralysis, repellent, food refusal and stomach toxicity action of contace poison, do not develop immunity to drugs, do not injure characteristics such as natural enemy.The plant polyphenol bacteriostatic agent is sprayed onto plant surface, has on the one hand diseases prevention, antibacterial and to the effect of insect resistance food, having water conservation on the other hand and regulate the effect of water balance. spray (dipping in) can also be under strong wind, dry climatic condition in plant surface simultaneously, regulate and control the nursery stock dehydration effectively, build up resistance.
Embodiment
Following embodiment illustrates the present invention rather than limits its protection domain.
The preparation of the composite bacteriostatic agent of embodiment 1 plant polyphenol
At first take by weighing commercially available powdery valonea extract of 2g and the commercially available licorice powder of 4g.Under room temperature and condition of normal pressure, 2g powdery valonea extract is added in 1 premium on currency, treat that it dissolves fully after, add the 4g licorice powder again, treat promptly to obtain the composite bacteriostatic agent of plant polyphenol of the present invention after it dissolves fully.
The preparation of the composite bacteriostatic agent of embodiment 2 plant polyphenols
At first take by weighing commercially available powdery valonea extract of 2g and the commercially available ginkgo leaf powder of 4g.Under room temperature and condition of normal pressure, 2g powdery valonea extract and 4g ginkgo leaf powder are added in 1 premium on currency simultaneously, treat promptly to obtain the composite bacteriostatic agent of plant polyphenol of the present invention after it dissolves fully.
The preparation of the composite bacteriostatic agent of embodiment 3 plant polyphenols
At first take by weighing commercially available powdery valonea extract of 2g and the commercially available scutelloside phenol of 4g.Then, under 40 ℃ of conditions with normal pressure, 4g scutelloside phenol is added in 1 premium on currency, treat that it dissolves fully after, add 2g powdery valonea extract again, treat promptly to obtain the composite bacteriostatic agent of plant polyphenol of the present invention after it dissolves fully.
Determining of the composite bacteriostatic agent minimum bactericidal concentrations of embodiment 4 plant polyphenols of the present invention (MIC)
With bacillus subtilis, Escherichia coli, bacillus megaterium and staphylococcus aureus is test strain, adopts the Oxford agar diffusion method, investigates the bactericidal effect of variable concentrations plant polyphenol bacteriostatic agent of the present invention, finds out minimum bacteriocidal concentration (MIC).
Preparation for the examination bacterial suspension: will be seeded on the test tube slant medium under the aseptic condition respectively for the examination bacterial classification, and put in 37 ℃ of constant incubators and cultivated 20 hours, treat that bacterium grows, add sterile distilled water, dissolving, vibration forms suspension bacteria liquid.
With sterile solid LB medium heating and melting, be cooled to 50~60 ℃, pour in the sterile petri dish, drip several suspension bacteria liquids of preparation as stated above, shaking up gently, the thickness of medium is 2~2.5mm.Aseptic Oxford cup is put on the flat board that respectively contains bacterium, each dull and stereotyped 4, implantation concentration is the plant polyphenol bacteriostatic agent aqueous solution 0.20ml that 0.25mg/ml, 0.5mg/ml, 1.0mg/ml, 1.5mg/ml, 2.0mg/ml, 2.5mg/ml, 3.0mg/ml, 5.0mg/ml prepare according to embodiment 1 mode respectively, cultivated 24 hours at 37 ℃, observe the sterilization conditions of each sample, find out minimum bactericidal concentration (MIC).It the results are shown in following table 1.
Table 1. variable concentrations plant polyphenol is composite
The bactericidal effect of bacteriostatic agent solution
Concentration mg/ml | 0.25 | 0.5 | 1 | 1.5 | 2 | 2.5 | 3 | 5 |
Bacillus subtilis escherichia coli and staphylococcus aureus bacillus megaterium | _ _ _ _ | _ _ _ _ | + + + + | + + + + | + + + + | + + + + | + + + + | + + + + |
Annotate :-expression does not have bactericidal properties; + expression has bactericidal properties
The result of table 1 shows that the minimum inhibitory concentration of plant polyphenol bacteriostatic agent of the present invention is 1mg/ml.
The composite bacteriostatic agent of embodiment 5 plant polyphenols of the present invention is to the bactericidal effect experiment of specified strain
Adopt the Oxford agar diffusion method, with the 0.5mg/ml minimum inhibitory concentration, checking the present invention's plant polyphenol bacteriostatic agent is to the bactericidal effect of 9 kinds of bacterial strains such as bacillus subtilis, Escherichia coli, bacillus megaterium, staphylococcus aureus, consor Dothiorella ribis, blight of chestnut bacterium, pears phyllosticta, black and white wheel branch spore, flax thorn dish spore.
Preparation for the examination bacterial suspension: will be seeded on the test tube slant medium under the aseptic condition respectively for the examination bacterial classification, and put in 37 ℃ of constant incubators and cultivated 20 hours, treat that bacterium grows, add sterile distilled water, dissolving, vibration forms suspension bacteria liquid.
With sterile solid LB medium heating and melting, be cooled to 50~60 ℃, pour in the sterile petri dish, drip several suspension bacteria liquids respectively by the method preparation of embodiment 4, shake up gently, the thickness of medium is 2~2.5mm.Aseptic Oxford cup is put on the flat board that respectively contains bacterium, each dull and stereotyped 4, implantation concentration is the present invention's of 0.5mg/ml plant polyphenol bacteriostatic agent aqueous solution 0.20ml respectively, cultivates 24 hours at 37 ℃, observe sterilization conditions, measure the inhibition zone size that every kind of bacterial strain produces.
The composite bacteriostatic agent of table 2. plant polyphenol of the present invention
Bactericidal effect to different strain
Bacterial strain | Inhibition zone size (mm) |
Bacillus subtilis Escherichia coli bacillus megaterium staphylococcus aureus consor Dothiorella ribis Cryphonectria Parasitica pears phyllosticta black and white wheel branch spore flax thorn dish spore | 17.5 17.0 16.5 17.5 18.0 18.0 18.0 18.0 8.0 |
The criterion of reference drug sensitive experiment is Gao Min at inhibition zone greater than 9 millimeters, and presentation of results the present invention's of table 2 plant polyphenol bacteriostatic agent is to all having the obvious sterilization effect for the examination bacterial classification.