CN101775022A - Ion-pair compound of chelerythrine from Chinese medicine extracts, preparation and application thereof - Google Patents
Ion-pair compound of chelerythrine from Chinese medicine extracts, preparation and application thereof Download PDFInfo
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Abstract
The invention relates to an ion-pair compound of chelerythrine from Chinese medicine extracts, a preparation method and application thereof. Organic acids of the invention mainly comprise chlorogenic acid, gallic acid, shikimic acid, oleanolic acid, ursolic acid, benzoic acid, salicylic acid, tartaric acid, lactic acid, sorbic acid, fumaric acid, malic acid, citric acid, maleic acid, glycyrrhizic acid, glycyrrhetinic acid, carnosic acid, P-hydroxycinnamic acid, rosmarinic acid, succinic acid, ferulic acid, protocatechuic acid, cinnamic acid, caffeic acid, chicoric acid or cortex pseudolaricis acetic acid. The ion-pair compound prepared from the chelerythrine and the organic acids has the antibacterial biological activity. The invention relates to the application of the ion-pair compound of the chelerythrine to medicine, animal remedy or daily use cosmetics.
Description
Technical field
The present invention relates to a kind of ion pair medicine and preparation method of the white chelerythrine from Chinese medical extract, and this medicine specifically is a pharmaceutical applications in people's medication, medication once and daily chemical products at antibiosis, particularly relate to the preparation that the ion-pair compound that contains white chelerythrine and organic acid generation is used for veterinary drug or cosmetics of everyday use, refer in particular to application at the veterinary drug and the medicated feed additive of the antibiotic direction of animal.
Background technology
In recent years, because human for antibiotic abuse, bacterium presents fast rise trend to all kinds of antibiotic resistances always.China is one of the most serious country of abuse of antibiotics in the world, and the bacterial drug resistance problem that causes thus is particularly outstanding.Clinical more isolating bacteriums are occupied first place in the world to the resistance of some drugs.For example, in the streptococcus aureus that infect in Shanghai, 80% has produced the resistance to penicillin G.The application of Cefotaxime, rocephin grade in an imperial examination triple-substituted cephalosporins is general day by day, and selecting for use of microbiotic kind is obviously leading.Quinolone antibiotic entered China only more than 20 year, and resistant rate has reached 60%～70%.Many common antibiotics antibacterial efficacies day by day lower, and antibiotics resistance has become the difficult problem that global public health faces.The World Health Organization (World Health Organization) claims that in the face of the antibiotics resistance problem that has increased rapidly since nearly half a century, medical circle is felt simply helpless.
In the herding industry, the drug-resistance of bacteria problem is equally also very serious.Modern breeding scale makes piglet lactation from 5-6 week shorten to 3-4 week, though at utmost brought into play the reproductive performance of sow, but produce some disadvantageous effects inevitably: wean back grice diarrhoea occurrence frequency obviously increases, piglet growth is obstructed, mortality ratio increases and extra medical expense strengthens, and has increased aquaculture cost.In order to prevent diarrhoea and to improve production performance, in the daily ration of be everlasting weanling pig and growing-finishing pig, add the germ resistance fodder additives, as microbiotic or microbiotic sample positive growth factor.Yet recent years, the public increased day by day to the tolerance of animal product pathogenic agent and the concern of antibiotic remains, forced people to go exploitation that the human consumer is safe from harm, and can play the substitute products of anti-microbial effect, the generation of prevention diarrhoea simultaneously again.
Before modern herbal medicine was born, herbal medicine was that the animal husbandry development of China has been made major contribution for thousands of years, and for example herbal medicine embodies distinctive effect at Chinese medicine aspect the sterilization of animal, anti-inflammatory.Based on traditional Chinese medical theory, most of heat-clearing and damp-drying drug class Chinese medicines all have (pressing down) bacterium activity extremely of varying degree, and have been widely used in veterinary applications, and are obtaining some achievements aspect the treatment infectation of bacteria.Because the singularity of Chinese medicine, bacterium is less to produce resistance to Chinese medicine, and that research has is efficient, wide spectrum, low chemical sproof Chinese medicine medicine, has strong reality significance for the treatment bacterial inflammation.Wherein, have alkaloid and organic acid anti-microbial activity as some by the ion-pair compound that alkaloid of forming from the herbal medicine natural component and organic acid generate, and alkaloid and organic acid may produce synergy, thereby strengthen the anti-microbial activity of ion-pair compound and enlarge antimicrobial spectrum, performance is similar to the prevention Animal diseases of Antibiotic Additive and promotes the growth effect in vivo.
White chelerythrine is the benzo phenanthridine alkaloid, has another name called toddaline, toddaline.Mainly be distributed in the plants such as papaveracease, Rutaceae, Berberidaceae.White chelerythrine mainly from bloodroot such as Pink Plumepoppy Herb, snowpoppy, pilewort etc. and rutaceae such as two-sided pin, Root of Asiatic Toddalia extraction separation obtain or by synthetic preparation.That its pharmacological action is mainly is antibiotic, anti-inflammatory, desinsection, kill maggot, improve liver function, anti-hepatic fibrosis, strengthening immunity, antitumor etc., because of it is paid much attention in the unique effect aspect the pharmacology.The latest find white chelerythrine can reduce the hepatic fibrosis rats model liver organization TGF-β 1 and the α-SMA of tetrachloro-methane induction, thus the protection liver.White chelerythrine also can suppress the change of high sugared inductive neonatal rat myocardial cell form and function, and can suppress expression and the activity of PKC-α, PKC-β 2, NF-κ B and C-fos, and the myocardial cell in the sugared environment of height is had provide protection.
White chelerythrine molecular formula: C
21H
18O
4N
+Molecular weight: 348.37, structural formula is as follows:
Extraction separation obtains or passes through in the research of the synthetic white chelerythrine for preparing in using from herbal medicine (bloodroot such as Pink Plumepoppy Herb, snowpoppy, pilewort etc. and rutaceae such as two-sided pin, Root of Asiatic Toddalia), except using single white chelerythrine composition to be used for clinical or herding is produced, the trend of use has also appearred white chelerythrine and other compositions are united at present.For example, mention in Chinese patent application 200610136957.4, " application in prevention and cure of snail fever of sanguinarine(e) or white chelerythrine " sanguinarine(e) and white chelerythrine are carried out the salinization processing respectively and mix the back to be used for prevention and cure of schistosomiasis, obtained good Synergistic treatment effect, salinization is simultaneously handled and has also been improved bioavailability of medicament.Yet aforesaid method concentrates on the two is mixed or single processing, is used for producing synergy by uniting to make, and the combination medicine that is obtained not is single medicine, and its physical and chemical stability and bioavailability are all not as the pure compound composition.
Chinese patent CN 100572380C discloses a kind of Berberine that extracts from the Chinese medicine coptis and another kind of traditional Chinese medicine ingredients houttuynine sodium bisulfite sodium salt generates a kind of Berberine ion-pair compound by combination reaction.Behind the prepared ion-pair compound of this method, enlarge the antimicrobial spectrum of Berberine, strengthened antibacterial effect, reduce the generation of Resistant strain, and increased the fat-soluble of Berberine,, thereby be expected to improve the clinical efficacy of Berberine for the bioavailability that increases Berberine provides possibility.Yet, this method must select two kinds of physico-chemical properties similar, and the identical macro-radical of pharmacological action carries out chemical combination, enter in the body back two kinds of molecules performance drug effect that dissociates respectively, mean that for the present invention must to select a kind of and white chelerythrine to have physico-chemical property similar, and the compound of the Chinese medical extract that pharmacological action is identical is handled, this has not only improved production cost, strengthened the reaction difficulty, must determine the primary and secondary/proportionlity of two kinds of medicinal ingredientss according to " monarch; minister; assistant; make " medicine relation of dividing of herbal medicine simultaneously, and must rely on numerous clinical experiments to be proved that can these two kinds of active pharmaceutical ingredientss produce synergistic effect, this has increased the difficulty of research and development white chelerythrine combination drug undoubtedly.
Comprehensively described, a kind of bioavailability that can improve white chelerythrine need be provided at present, enlarge white chelerythrine antimicrobial spectrum, strengthen the white chelerythrine drug effect, introduce the method for other medicinal ingredientss of Cheap highly effective and the product of white chelerythrine combination drug thereof simultaneously.
Summary of the invention
The inventor is based on by for many years to from herbal medicine (bloodroot such as Pink Plumepoppy Herb, snowpoppy, pilewort etc. and rutaceae such as two-sided pin, Root of Asiatic Toddalia) white chelerythrine of extract research accumulation, breakthrough is for the conventional thought of white chelerythrine exploitation, promptly no longer be confined to select to be suitable for the suitable carrier or the compatibility agent of white chelerythrine, also no longer in selecting physico-chemical property similar to white chelerythrine, and other identical medicinal compounds of pharmacological action are handled, but a kind of antibacterial synergistic effect that can either strengthen white chelerythrine of creationary selection, can significantly improve the Chinese medicine single component of bioavailability again, belong to Cheap highly effective simultaneously and easily carry out the simple compounds of combination reaction, and the two is combined into new pairing compound (being ion-pair compound) by intermolecular electrostatic attraction effect unconventionally, thereby finally obtain a kind of new ion-pair compound from Chinese medicine.Briefly, be that the contriver passes through a large amount of creative works, it is unexpected that discovery can select some and white chelerythrine to have the organic acid group of identical bactericidal effect, itself and white chelerythrine are carried out combination reaction, thereby acquisition can realize the ion-pair compound of the white chelerythrine composite organic acid of the object of the invention.On the other hand, the present invention by a large amount of experimental selection the multiple organic acid that is suitable for the object of the invention.
Therefore, it is that a kind of ion-pair compound that derives from the white chelerythrine of herbal medicine (bloodroot such as Pink Plumepoppy Herb, snowpoppy, pilewort etc. and rutaceae such as two-sided pin, Root of Asiatic Toddalia) or chemical combination generation is provided first that water is invented first purpose, it has the structure shown in the general formula I
Wherein, organic acid RCOO
-Be selected from chlorogenic acid, gallic acid, shikimic acid, Oleanolic Acid, ursolic acid, phenylformic acid, Whitfield's ointment, tartrate, lactic acid, Sorbic Acid, fumaric acid, oxysuccinic acid, citric acid, toxilic acid, Potenlini, glycyrrhetinic acid, Salvin, Hydroxycinnamic acid, rosmarinic acid, succsinic acid, forulic acid, Protocatechuic Acid, styracin, coffic acid, chicoric acid or Pseudolarix acid B with anti-microbial activity.
In one embodiment, described organic acid RCOO
-Be selected from chlorogenic acid, gallic acid, shikimic acid, Oleanolic Acid, Salvin, rosmarinic acid, phenylformic acid, Whitfield's ointment.
Second purpose of the present invention provides the method for the above-mentioned simplification compound of preparation (being ion-pair compound), comprises according to following reaction formula being prepared described ion-pair compound:
After wherein chelerythrine alkali salt and organic acid sodium salt react completely, produce precipitation, sedimentation and filtration, drying obtain described ion-pair compound, the hydrochloride that the white chelerythrine hydrochloride is made from the white chelerythrine of the white chelerythrine of Chinese medical extract or chemosynthesis.
In one embodiment, in the above-mentioned reaction chelerythrine alkali salt and organic acid sodium salt are made the aqueous solution or saturated aqueous solution respectively under 60 ℃-100 ℃, mix again after the filtration and produce precipitation, filter, drying precipitatedly obtain described ion-pair compound, the ion-pair compound of gained is carried out recrystallization purifying by organic solvent.
The 3rd purpose of the present invention provides the method for the another kind of above-mentioned simplification compound of preparation (being ion-pair compound), comprising:
(1) with the white chelerythrine hydrochloride aqueous solution and organic acid sodium salt aqueous solution, produce precipitation, precipitation after filtration, dry ion-pair compound;
(2) will be derived from the white chelerythrine of Chinese medical extract or the white chelerythrine of chemosynthesis and be dissolved in the organic solvent, and add organic acid then and stir until reacting completely, concentrate drying gets ion-pair compound;
(3) white chelerythrine hydrochloride, organic acid sodium salt are dissolved in the water respectively, then with two kinds of solution by etc. mol ratio carry out mixing, stir and to be precipitated in a large number, precipitation after filtration, dry ion-pair compound; Maybe will be derived from the white chelerythrine of Chinese medical extract or the white chelerythrine of chemosynthesis is dissolved in the organic solvent, and organic acid is dissolved in the organic solvent, then with two kinds of solution by etc. mol ratio mix, stir until reacting completely, concentrate drying gets ion-pair compound;
Above-mentioned organic solvent is selected from methyl alcohol, ethanol, acetone, ethyl acetate or chloroform solvent.
In one embodiment, aforesaid method also comprises the ion-pair compound of gained is heavily tied the product purifying by organic solvent.
In one embodiment, above-mentioned reaction can also be that the chelerythrine alkali salt is hydrochloride, hydrosulfate, nitrate or phosphoric acid salt; Described organic acid sodium salt is by organic acid and NaOH, Na
2CO
3Or NaHCO
3The reaction gained.
The 4th purpose of the present invention provides the purposes of ion-pair compound in the preparation antibacterial-anti-inflammatory drug of the prepared white chelerythrine of a kind of aforesaid method, and wherein in one embodiment, described medicine is people's medication or veterinary medicine.
The 5th purpose of the present invention provides the degerming or the anti-inflammatory daily chemical products of the ion-pair compound that contains above-mentioned white chelerythrine, wherein in one embodiment, described daily chemical products is makeup, articles for washing, oral-cavity article or clean body articles for use, flavouring agent, reodorant, expelling parasite go out harmful product or other daily chemical products etc.
The 6th purpose of the present invention provides the fodder additives or the medicated feed additive of the ion-pair compound that contains above-mentioned white chelerythrine, wherein in one embodiment, described fodder additives or medicated feed additive are poultry, livestock or aquatic feeds or fodder additives.
Definition:
Unless specialize, " ion-pair compound " of the present invention, or " ion pair medicine ", be meant the water-insoluble compound that macromolecule organic acid and organic bases generate by intermolecular electrostatic attraction combination, possess hydrophilic property is little, the characteristics that lipotropy is big, its spectrum are also before different with the formation ion-pair compound.The formation of ion-pair compound has changed physico-chemical properties such as monomer molecule solvability, stability, and can bring into play activity separately, might produce synergy, thereby influence curative effect of medication; Simultaneously be easy to penetrate lipophilic cytolemma again, improve absorption.
" organic acid " is meant that some have the tart organic compound.Modal organic acid is a carboxylic acid, and its acidity comes from carboxyl (COOH).Unless specialize, " organic acid " of the present invention, having another name called " organic acid with anti-microbial activity " is some medium-height grass the effective elements of the medicine, that majority has is antimycotic, antibacterium, biological activity such as antiviral.Organic acid has different effects to the micro-flora of animal intestines and stomach, because mainly the appropriate pH value environment of harmful microorganism growth mostly is neutral, the growth and breeding in sour environment and beneficial microorganisms such as lactobacillus suit, organic acid reaches the effect of the harmful germ quantity of indirect reduction by the pH value that reduces gastrointestinal tract environment, in addition, organic acid can also reach anti-microbial effect by pH value and the gathering of poisonous negatively charged ion in bacterium in the cytolemma that destroys germ, the basal metabolism function that suppresses bacterium, the adjusting bacterium.Because pH value, concentration are different with the state that dissociates, different organic acids has different antibacterial effects.
Following table 1 has been listed organic acid structural formula with anti-microbial activity related among the present invention
Table 1 organic acid structural formula
" degerming or anti-inflammatory daily chemical products ", makeup, articles for washing, oral-cavity article or clean body articles for use, flavouring agent, reodorant, the expelling parasite that refers to contain ion pair medicine of the present invention go out harmful product or other daily chemical products.Common daily chemical products comprises: cosmetics, cleaning makeup, skin care product, hair cosmetic, medicated soap or perfumed soap or Liquid soap, washing composition or washings or washing powder, thimerosal or sterilizing agent, Liquid soap or scale remover, toothpaste, collutory, shoe polish, floor wax etc.
" fodder additives or the medicated feed additive that contain the ion-pair compound of above-mentioned white chelerythrine " refers to be used to prevent the non-nutritional feed additive of Animal diseases, this class additive is to the nonnutritive effect of animal, but can be used for reaching the promotion growth of animal, improve the price of deed by preventing and treating eqpidemic disease, the absorption of promotion animal digestion etc., reduce feed cost, obtain the more purpose of large economy benefit.Common " fodder additives that contains the ion-pair compound of above-mentioned white chelerythrine " comprising: antimicrobial growth promoter (microbiotic, antiseptic-germicide etc.), hormones additive, enzyme additive, microbe additive, anthelmintic drug additive, Chinese medicine additive, calm class additive etc.
Beneficial effect
1. the prepared white chelerythrine ion-pair compound possess hydrophilic property of the present invention is little, and the characteristics that lipotropy is big have changed monomer molecule solvability physico-chemical property, are easy to penetrate lipophilic cytolemma again, improves to absorb;
2. prepared white chelerythrine ion-pair compound both can have been brought into play active separately, the synergy of generation is also arranged, thereby improved bioavailability, further promoted curative effect of medication;
3. the present invention utilizes white chelerythrine and chlorogenic acid, and the ion-pair compound that organic acids such as gallic acid, phenylformic acid generate has increased the stability of white chelerythrine greatly, helps using in prolonged preservation and the body;
4. by anti-microbial activity mensuration, bioavailability study and the toxicologic study of white chelerythrine ion-pair compound are found, this ion-pair compound can strengthen the white chelerythrine antibacterial effect, improve the bioavailability of white chelerythrine, and, reduce the generation of Resistant strain for the antimicrobial spectrum that enlarges white chelerythrine provides possibility;
Embodiment
As previously mentioned, according to the disclosed content in front, one of skill in the art can use the present invention to greatest extent.Should be pointed out that the present invention aims to provide a kind of new antibacterial-anti-inflammatory drug.Following embodiment partly provide be enough to confirm the curative effect of described medicine after, those skilled in the art are according to the routine techniques of pharmaceutical applications, daily chemical products purposes and feed purposes, can expect white chelerythrine ion-pair compound of the present invention fully can be used in above-mentioned field and can realize expectation function, therefore the following examples can help those skilled in the art more fully to understand the present invention, but do not limit the present invention in any way.
White chelerythrine preparation technology: with Herba Macleayae Cordatae extract or other plant milk extracts that contains white chelerythrine is raw material, add suitable quantity of water, be heated to 60～90 ℃ of dissolvings, filter, in the macroporous adsorption resin chromatography post of filtrate flow through installing, use Different concentrations of alcohol-aqueous solution gradient elution more respectively, collect the elutriant that mainly contains white chelerythrine respectively, carry out crystallization under proper condition with ethanol or methyl alcohol again, the crystal heating for dissolving is in methyl alcohol or ethanol, add hydrochloric acid or sulfuric acid and white chelerythrine salify and generate white chelerythrine hydrochloride or white chelerythrine hydrosulfate, the hydrochloride of gained white chelerythrine or the purity of hydrosulfate reach (HPLC) more than 98%.
Embodiment 1: the preparation method of the ion-pair compound that white chelerythrine hydrochloride and shikimic acid sodium salt generate
10.0g is dissolved in the 1000mL water with white chelerythrine hydrochloride (being provided by Hunan Engineering Research Center of Botanical Extract), is heated to more than 90 ℃, all dissolves until the white chelerythrine hydrochloride, makes the white chelerythrine hydrochloride aqueous solution (A); Shikimic acid sodium salt 1.0g is dissolved in is mixed with shikimic acid sodium-salt aqueous solution (B) in the 200mL water;
Under the condition (B) slowly joined in (A) more than 90 ℃, 90 ℃ are constantly stirred, and reaction 30min is cooled to room temperature, filters, and with pure water thorough washing precipitation, precipitates 60 ℃ of vacuum-dryings, promptly gets flaxen crystal, carries out
1H-NMR,
13The white chelerythrine shikimic acid ion-pair compound that the compound that C-NMR, MS analysis obtains is.
Embodiment 2: the preparation method of the ion-pair compound that white chelerythrine and Salvin generate
10.0g is dissolved in the 2000mL acetone with white chelerythrine (being provided by Hunan Engineering Research Center of Botanical Extract), stirring and dissolving.Directly add Salvin 10.0g then, normal temperature stirs 45min down, concentrates, and drying obtains solid substance, obtains light yellow crystal with ethyl acetate is refining again, carries out
1H-NMR,
13It is white chelerythrine Salvin ion-pair compound that C-NMR, MS analyze the compound that obtains.
Embodiment 3: the preparation method of the ion-pair compound that white chelerythrine hydrochloride and rosmarinic acid sodium generate
White chelerythrine hydrochloride (providing by Hunan Engineering Research Center of Botanical Extract) 4.5g is provided is dissolved in the 1L water, add 4.0g rosmarinic acid sodium then, stir 30min in 80 ℃, after the cooling, filter washing, drying further obtains light yellow crystal with refining methanol again.By carrying out
1H-NMR,
13It is the ion-pair compound of white chelerythrine rosmarinic acid that C-NMR, MS analyze the compound that obtains.
Embodiment 4: the preparation method of the ion-pair compound that white chelerythrine and chlorogenic acid generate
Getting white chelerythrine. (being provided by Hunan Engineering Research Center of Botanical Extract) 3.5g is dissolved in the methanol solvate, directly adds the 3.5g chlorogenic acid again.Stirring at normal temperature 30min concentrates, and drying is more further with the refining light yellow crystal that obtains of ethanol.By carrying out
1H-NMR,
13It is the ion-pair compound of white chelerythrine chlorogenic acid that C-NMR, MS analyze the compound that obtains.
Embodiment 5: the preparation method of the ion-pair compound that white chelerythrine hydrochloride and Sodium Benzoate generate
White chelerythrine hydrochloride (providing by Hunan Engineering Research Center of Botanical Extract) 4.5g is provided is dissolved in the 1L water, add the 1.5g Sodium Benzoate then, stir 30min in 80 ℃, after the cooling, filter washing precipitation, drying further obtains light yellow crystal with refining methanol again.By carrying out
1H-NMR,
13It is the ion-pair compound of white chelerythrine chlorogenic acid that C-NMR, MS analyze the compound that obtains.
Embodiment 6: the preparation method of the ion-pair compound that white chelerythrine and gallic acid generate
White chelerythrine (being provided by Hunan Engineering Research Center of Botanical Extract) 3.5g is provided is dissolved in the acetone solvent, get gallic acid 2.0g again and be dissolved in the acetone, both solution are mixed, normal temperature stirs 30min down.Concentrate, drying, with the gained precipitation with ethyl acetate refining light yellow crystal.By carrying out
1H-NMR,
13It is the ion-pair compound of white chelerythrine gallic acid that C-NMR, MS analyze the compound that obtains.
Embodiment 7: the preparation method of the ion-pair compound that white chelerythrine and Whitfield's ointment generate
Get among the Whitfield's ointment 1.4g dissolving ethyl acetate 100ml .3.5g white chelerythrine (being provided by Hunan Engineering Research Center of Botanical Extract) is provided then, normal temperature stirs 45min down, concentrates, and drying is with the light yellow crystal of gained precipitation with refining methanol.By carrying out
1H-NMR,
13It is the salicylic ion-pair compound of white chelerythrine that C-NMR, MS analyze the compound that obtains
Embodiment 8: the antibacterial activity in vitro of white chelerythrine ion-pair compound is measured
The ion-pair compound and the white chelerythrine of the ion-pair compound generation of ion-pair compound, white chelerythrine and rosmarinic acid generation that the ion-pair compound that the ion-pair compound that the ion-pair compound that above-mentioned prepared white chelerythrine and chlorogenic acid are generated, white chelerythrine and Whitfield's ointment generate, white chelerythrine and phenylformic acid generate, white chelerythrine and gallic acid generate carry out bacteriostatic activity mensuration.
Bacterial classification is Hunan Inst. of Microbe's isolation identification gained, and bacterial classification is respectively: intestinal bacteria, the configuration concentration of streptococcus aureus test sample are 2000 μ g/mL; Establish corresponding dissolved matrix contrast in the antibiotic process; Adopt agar plate diffusion process and tube dilution method to measure.
MIC measuring method: get 13 of sterile test tube, be arranged in a row, remove the 1st pipe and add the bacterium liquid of 1.8mLMH with 1000 times of meat soup dilutions, all the other every pipes add MH meat soup 1mL, add antibacterials stoste 0.2mL mixing at the 1st pipe, draw 1mL to the 2 pipes then, draw 1mL to the 3 pipes behind the mixing again, doubling dilution to the 12 is managed so continuously, and absorption 1mL discards from the 12nd pipe, and the 13rd pipe is not for containing the growth control of medicine.Cultivating 24h for 37 ℃, is minimal inhibitory concentration (MIC) with the minimum concentration of no bacterial growth.
Table 2 in-vitro antibacterial experimental result n=6
Annotate: B: white chelerythrine; BL: the ion-pair compound of white chelerythrine chlorogenic acid; BS: the salicylic ion-pair compound of white chelerythrine; BB: the benzoic ion-pair compound of white chelerythrine; BM: the ion-pair compound of white chelerythrine gallic acid; BD: the ion-pair compound of white chelerythrine rosmarinic acid.
Through the antibacterial activity in vitro determination experiment, above-mentioned research white chelerythrine ion-pair compound has stronger bacteriostatic action to intestinal bacteria, streptococcus aureus, its MIC all is lower than white chelerythrine, synergy owing to white chelerythrine and organic acid existence is described, has strengthened the antibacterial effect of the ion-pair compound of white chelerythrine and organic acid generation.
Embodiment 9: the bioavailability test of white chelerythrine ion-pair compound
Adopt HPLC-UV method research white chelerythrine, the ion-pair compound that white chelerythrine and chlorogenic acid generate, the ion-pair compound that white chelerythrine and Whitfield's ointment generate, the ion-pair compound that white chelerythrine and phenylformic acid generate, the ion-pair compound that white chelerythrine and gallic acid generate, the ion-pair compound that the ion-pair compound that white chelerythrine and rosmarinic acid generate generates totally 5 kinds of ion-pair compounds to irritate the Plasma Concentration after the administration of stomach mode, compare pharmacokinetic parameter, to disclose the medicine rule and the bioavailability thereof of dynamic change in animal body.
Instrument and reagent: Agilent 1100 high performance liquid chromatographs, Quat Pump pump, VWD ultraviolet-visible detector, the full-automatic sampler of Agilent, Agilent chromatographic working station.Pipettor, GILSON; FZQ-2 type turbine mixer (Tai County, Jiangsu medical apparatus and instruments factory); TGL-16G supercentrifuge (Anting Scientific Instrument Factory, Shanghai).Methyl alcohol, acetonitrile, Glacial acetic acid are chromatographically pure, and hydrochloric acid, ethyl acetate are analytical pure, and water is sub-boiling distillation water.
Laboratory animal: the Beagle dog that grows up, body weight (9kg ± 0.6kg), male and female half and half.
Chromatographic condition chromatographic column: anti-phase C18 post: Lichrospher C R (4.6mm * 250mm, 5 μ m); Moving phase: the glacial acetic acid aqueous solution of acetonitrile-1% (10: 90); Flow velocity: 1.0mLmin
-1Column temperature: 30 ℃; Wavelength: 284nm; Sample size: 20 μ L.
Plasma sample is handled: get plasma sample 0.5mL, add 1% Glacial acetic acid 50mL, the vortex mixing adds (3molL again
-1) hydrochloric acid soln 300mL, limit edged vortex adds ethyl acetate extraction 4 times, each 2mL, and the separating ethyl acetate layer merges, and N2 dries up under 37 ℃ of water-baths, and residue dissolves with methyl alcohol 200mL, and centrifugal, it is standby to get supernatant liquor.Sample introduction 20 μ L, the record peak area adopts the external standard calibration curve method quantitative with peak area ratio.
Sample collecting and processing: gather blank blood sample before the administration, the administration group is in 0,0.25,0.5,1.0,1.5,2.0,4.0,5.0,7.0,9.0,12.0, and 24.0h gets blood 5mL at veins of lower extremity, the blood sample that collects is put contain in the heparin centrifuge tube centrifugal (2000rmin
-1) 10min, get upper plasma-20 ℃ storage.
The calculating of pharmacokinetic parameter: DAS software The Fitting Calculation, determine compartment model, main pharmacokinetic parameters sees Table 3.
Table 3Beagle dog is irritated the main pharmacokinetic parameter behind the stomach medicine
Annotate: B: white chelerythrine; BL: the ion-pair compound of white chelerythrine chlorogenic acid; BS: the salicylic ion-pair compound of white chelerythrine; BB: the benzoic ion-pair compound of white chelerythrine; BM: the ion-pair compound of white chelerythrine gallic acid; BD: the ion-pair compound of white chelerythrine rosmarinic acid.
The result shows that white chelerythrine ion-pair compound group and white chelerythrine group compare, and Tmax shortens, and Cmax improves, and bioavailability improves greatly.
Embodiment 10: white chelerythrine and shikimic acid ion-pair compound are for the effect of domestic animal dysentery characterized by white mucous stool disease
According to method with embodiment 1, the white chelerythrine of preparation and shikimic acid ion-pair compound.
Weight ratio by 0.05% is mixed with the shikimic acid ion-pair compound white chelerythrine with feed, be used to feed 10 piglets (wherein 6 are the dysentery characterized by white mucous stool piglet), adds up the piglet number of the appearance dysentery characterized by white mucous stool in 5 days, obtains table 4.
Table 4 medicine is to the therapeutic action of domestic animal dysentery characterized by white mucous stool disease
As shown in Table 4, the piglet of treatment group no longer includes the dysentery characterized by white mucous stool piglet basically after 3 days, and the dysentery characterized by white mucous stool symptom of piglet is significantly improved.By contrast, the healthy piglet of control group all suffered from dysentery characterized by white mucous stool disease at the 4th day.
Embodiment 11: white chelerythrine and shikimic acid ion-pair compound are for the effect of poultry dysentery characterized by white mucous stool disease
According to method with embodiment 1, the white chelerythrine of preparation and shikimic acid ion-pair compound.
Fall white chelerythrine by 0.01% weight ratio and mix with feed, be used to feed 20 commodity Dai Luoman laying hens (wherein 10 are the dysentery characterized by white mucous stool chicken), add up the chicken number of the appearance dysentery characterized by white mucous stool in 5 days, obtain table 5 with the shikimic acid ion-pair compound.
Table 5 medicine is to the therapeutic action of poultry dysentery characterized by white mucous stool disease
As shown in Table 5, treatment group no longer includes the dysentery characterized by white mucous stool chicken basically after 3 days, and the dysentery characterized by white mucous stool symptom is significantly improved.By contrast, the control group healthy chicken was all suffered from dysentery characterized by white mucous stool disease at the 4th day.
Comprehensively described, white chelerythrine ion-pair compound possess hydrophilic property of the present invention is little, and the characteristics that lipotropy is big have changed physico-chemical properties such as white chelerythrine monomer molecule solvability, stability; Measure by the anti-microbial activity to white chelerythrine, organic acid and this ion-pair compound, find that there are synergy in white chelerythrine and organic acid, the antibacterial effect of the ion-pair compound that both generate obviously strengthens; In addition, by to white chelerythrine and this ion-pair compound bioavailability study, find that the bioavailability of organic acid and white chelerythrine generation ion-pair compound obviously increases, thereby can improve the clinical efficacy of white chelerythrine effectively.Therefore, white chelerythrine ion-pair compound of the present invention has wide pharmaceutical applications prospect, and can be used in the production of animal-feed, daily chemical products.
Claims (10)
1. the ion-pair compound of a white chelerythrine, it has the structure shown in the general formula I,
Wherein, organic acid RCOO
-Be selected from chlorogenic acid, gallic acid, shikimic acid, Oleanolic Acid, ursolic acid, phenylformic acid, Whitfield's ointment, tartrate, lactic acid, Sorbic Acid, fumaric acid, oxysuccinic acid, citric acid, toxilic acid, Potenlini, glycyrrhetinic acid, Salvin, Hydroxycinnamic acid, rosmarinic acid, succsinic acid, forulic acid, Protocatechuic Acid, styracin, coffic acid, chicoric acid or Pseudolarix acid B with anti-microbial activity.
2. ion-pair compound according to claim 1, wherein said organic acid RCOO
-Be selected from chlorogenic acid, gallic acid, shikimic acid, Oleanolic Acid, Salvin, rosmarinic acid, phenylformic acid or Whitfield's ointment.
3. the preparation method of claim 1 or 2 described ion-pair compounds comprises by following reaction being prepared the described ion-pair compound that obtains:
After wherein chelerythrine alkali salt and organic acid sodium salt react completely, produce precipitation, sedimentation and filtration, drying obtain described ion-pair compound; The hydrochloride that the white chelerythrine hydrochloride is made from the white chelerythrine of the white chelerythrine of Chinese medical extract or chemosynthesis; Organic acid in the described organic acid sodium salt is selected from chlorogenic acid, gallic acid, shikimic acid, Oleanolic Acid, ursolic acid, phenylformic acid, Whitfield's ointment, tartrate, lactic acid, Sorbic Acid, fumaric acid, oxysuccinic acid, citric acid, toxilic acid, Potenlini, glycyrrhetinic acid, Salvin, Hydroxycinnamic acid, rosmarinic acid, succsinic acid, forulic acid, Protocatechuic Acid, styracin, coffic acid, chicoric acid or the Pseudolarix acid B with anti-microbial activity.
4. method according to claim 3, in the wherein above-mentioned reaction chelerythrine alkali salt and organic acid sodium salt are made the aqueous solution or saturated aqueous solution respectively under 60 ℃-100 ℃, mix again after the filtration and produce precipitation, filter, drying precipitatedly obtain described ion-pair compound, the ion-pair compound of gained is carried out recrystallization purifying by organic solvent.
5. according to the preparation method of each described ion-pair compound of claim 3, obtain by any mode prepared in reaction in following three kinds of modes:
(1) with white chelerythrine salt brine solution and organic acid sodium salt aqueous solution, produce precipitation, precipitation after filtration, dry ion-pair compound;
(2) will be derived from the white chelerythrine of Chinese medical extract or the white chelerythrine of chemosynthesis and be dissolved in the organic solvent, and add organic acid then and stir until reacting completely, concentrate drying gets ion-pair compound;
(3) chelerythrine alkali salt, organic acid sodium salt are dissolved in the water respectively, then with two kinds of solution by etc. mol ratio carry out mixing, stir and to be precipitated in a large number, precipitation after filtration, dry ion-pair compound; Maybe will be derived from the white chelerythrine of Chinese medical extract or the white chelerythrine of chemosynthesis is dissolved in the organic solvent, and organic acid is dissolved in the organic solvent, then with two kinds of solution by etc. mol ratio mix, stir until reacting completely, concentrate drying gets ion-pair compound;
Above-mentioned organic solvent is selected from methyl alcohol, ethanol, acetone, ethyl acetate or chloroform solvent.
6. method according to claim 5 comprises that also the ion-pair compound with gained carries out recrystallization purifying by organic solvent.
7. according to each described method of claim 3-6, described is that the chelerythrine alkali salt is hydrochloride, hydrosulfate, nitrate or phosphoric acid salt; Described organic acid sodium salt is by organic acid and NaOH, Na
2CO
3Or NaHCO
3The reaction gained.
8. claim 1 or 2 described ion-pair compounds, or the prepared purposes of ion-pair compound in the preparation antibacterial-anti-inflammatory drug of claim 3-7, wherein said medicine is people's medication or veterinary medicine.
9. the degerming or the anti-inflammatory daily chemical products that contain any described ion-pair compound of claim 1-7.
10. contain any described ion-pair compound of claim 1-7 fodder additives or medicated feed additive, wherein said fodder additives or medicated feed additive are poultry, livestock or feeding additive aquatic animal or medicated feed additive.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103749981A (en) * | 2014-01-13 | 2014-04-30 | 江苏省农业科学院 | Application of protocatechuic acid as feed additive in livestock breeding |
| CN106138054A (en) * | 2015-04-28 | 2016-11-23 | 沈阳伟嘉牧业技术有限公司 | Chelerythrine application in treatment fowl drug resistance colibacillosis |
| CN113549085A (en) * | 2021-07-16 | 2021-10-26 | 湖北科技学院 | Synthetic method of natural product 6-HHC |
| CN114939068A (en) * | 2022-06-28 | 2022-08-26 | 四川大学 | Amorphous calcium phosphate composite material, preparation method thereof and application thereof in oral cavity |
-
2010
- 2010-02-09 CN CN201010110411A patent/CN101775022A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103749981A (en) * | 2014-01-13 | 2014-04-30 | 江苏省农业科学院 | Application of protocatechuic acid as feed additive in livestock breeding |
| CN106138054A (en) * | 2015-04-28 | 2016-11-23 | 沈阳伟嘉牧业技术有限公司 | Chelerythrine application in treatment fowl drug resistance colibacillosis |
| CN113549085A (en) * | 2021-07-16 | 2021-10-26 | 湖北科技学院 | Synthetic method of natural product 6-HHC |
| CN114939068A (en) * | 2022-06-28 | 2022-08-26 | 四川大学 | Amorphous calcium phosphate composite material, preparation method thereof and application thereof in oral cavity |
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