CN103520323A - Application of Nauclea officinalis extract to preparation of medicament for resisting methicillin-resistant Staphylococcus aureus - Google Patents
Application of Nauclea officinalis extract to preparation of medicament for resisting methicillin-resistant Staphylococcus aureus Download PDFInfo
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- CN103520323A CN103520323A CN201310473042.2A CN201310473042A CN103520323A CN 103520323 A CN103520323 A CN 103520323A CN 201310473042 A CN201310473042 A CN 201310473042A CN 103520323 A CN103520323 A CN 103520323A
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- radix naucleae
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- 239000000284 extract Substances 0.000 title claims abstract description 136
- 241000191967 Staphylococcus aureus Species 0.000 title claims abstract description 50
- RJQXTJLFIWVMTO-TYNCELHUSA-N Methicillin Chemical compound COC1=CC=CC(OC)=C1C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 RJQXTJLFIWVMTO-TYNCELHUSA-N 0.000 title claims abstract description 49
- 229960003085 meticillin Drugs 0.000 title claims abstract description 49
- 239000003814 drug Substances 0.000 title claims abstract description 24
- 238000002360 preparation method Methods 0.000 title abstract description 14
- 241001620257 Nauclea officinalis Species 0.000 title abstract description 12
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 58
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- 238000000034 method Methods 0.000 claims description 7
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- 238000012360 testing method Methods 0.000 description 30
- 241000894006 Bacteria Species 0.000 description 21
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- MYPYJXKWCTUITO-LYRMYLQWSA-N vancomycin Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C2C=C3C=C1OC1=CC=C(C=C1Cl)[C@@H](O)[C@H](C(N[C@@H](CC(N)=O)C(=O)N[C@H]3C(=O)N[C@H]1C(=O)N[C@H](C(N[C@@H](C3=CC(O)=CC(O)=C3C=3C(O)=CC=C1C=3)C(O)=O)=O)[C@H](O)C1=CC=C(C(=C1)Cl)O2)=O)NC(=O)[C@@H](CC(C)C)NC)[C@H]1C[C@](C)(N)[C@H](O)[C@H](C)O1 MYPYJXKWCTUITO-LYRMYLQWSA-N 0.000 description 17
- MYPYJXKWCTUITO-UHFFFAOYSA-N vancomycin Natural products O1C(C(=C2)Cl)=CC=C2C(O)C(C(NC(C2=CC(O)=CC(O)=C2C=2C(O)=CC=C3C=2)C(O)=O)=O)NC(=O)C3NC(=O)C2NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(CC(C)C)NC)C(O)C(C=C3Cl)=CC=C3OC3=CC2=CC1=C3OC1OC(CO)C(O)C(O)C1OC1CC(C)(N)C(O)C(C)O1 MYPYJXKWCTUITO-UHFFFAOYSA-N 0.000 description 17
- 230000002401 inhibitory effect Effects 0.000 description 16
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- 229960000479 ceftriaxone sodium Drugs 0.000 description 15
- FDRNWTJTHBSPMW-GNXCPKRQSA-L disodium;(6r,7r)-7-[[(2e)-2-(2-amino-1,3-thiazol-4-yl)-2-methoxyiminoacetyl]amino]-3-[(2-methyl-6-oxido-5-oxo-1,2,4-triazin-3-yl)sulfanylmethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate Chemical compound [Na+].[Na+].S([C@@H]1[C@@H](C(N1C=1C([O-])=O)=O)NC(=O)/C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NC(=O)C([O-])=NN1C FDRNWTJTHBSPMW-GNXCPKRQSA-L 0.000 description 15
- 229960003276 erythromycin Drugs 0.000 description 15
- 239000003112 inhibitor Substances 0.000 description 15
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- 238000000338 in vitro Methods 0.000 description 10
- 238000007619 statistical method Methods 0.000 description 10
- 206010041925 Staphylococcal infections Diseases 0.000 description 7
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- 238000004500 asepsis Methods 0.000 description 5
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- 239000011248 coating agent Substances 0.000 description 5
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- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
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- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The invention discloses application of water extract, ethanol extract, ethanol aqueous solution extract of Nauclea officinalis and corresponding petroleum ether abstract, ethyl acetate abstract, n-butanol abstract and water abstract of Nauclea officinalis to preparation of medicaments for resisting methicillin-resistant Staphylococcus aureus (MRSA). At the same time, the invention discloses a preparation method of the above extracts or abstracts. The experimental results show that water extract, ethanol extract, ethanol aqueous solution extract of Nauclea officinalis and corresponding ethyl acetate abstract, n-butyl alcohol abstract and water abstract have strong effect in resisting MRSA, and an ethyl acetate extraction position of 70% ethanol extract of Nauclea officinalis has the highest activity.
Description
Technical field
The invention belongs to field of traditional Chinese medicine pharmacy, relate to the new purposes of Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) extract.
Background technology
As everyone knows, current bacterial resistance has become the great public health problem of global range, and antibiotic abuse is the major reason that causes bacterial resistance.Methicillin-resistant staphylococcus aureus (Methicillin-resistant Staphylococcus aureus, hereinafter to be referred as " MRSA ") there is multi-drug resistant and with high incidence and high mortality, it is the important pathogenic bacteria that causes the diseases such as necrotizing pneumonia, serious septicemia, necrotizing fasciitis, be one of important pathogen of hospital infection and community's infection, to clinical treatment, brought great difficulty.MRSA is to most antimicrobial drug drug resistances, even the most effective vancomycin of current clinical antimicrobial agent is also started to occur drug resistance, once MRSA develops into the degree to the general drug resistance of vancomycin, the patient who infects MRSA will face the desperate situation pasting medical help.Therefore, the situation that directed toward bacteria drug resistance is on the rise, the antibacterials that find, exploitation have antimicrobial agent MRSA effect are very urgent.
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is Rubiaceae Nauclea plant, another name ebony, the yellow wood of spire, Mongolian gazelle wood, mountain Fel Ursi, Fel Ursi tree.Lignum et Radix Naucleae was medical material among the people originally, begin to be loaded in Guangzhou army and compile the books such as " conventional Chinese herbal medicine handbook " and " national Chinese herbal medicine compilation ", by 1977 version " Chinese Pharmacopoeia " record.Stem and the Pi Junneng of Lignum et Radix Naucleae are used as medicine, and have the effect of heat-clearing and toxic substances removing, reducing swelling and alleviating pain, are usually used in the treatment of the diseases such as cold, fever, laryngopharynx swelling and pain, acute conjunctivitis, acute tonsillitis, pneumonia, enteritis, dysentery.There is no at present the medicinal application of bibliographical information Lignum et Radix Naucleae methicillin-resistant staphylococcus aureus resistance.
Summary of the invention
The object of the invention is to develop the new purposes of Lignum et Radix Naucleae on pharmaceutical field.
One of object is, by Lignum et Radix Naucleae water or ethanol or ethanol water solution extract for the preparation of methicillin-resistant staphylococcus aureus resistance medicine.
Two of object is further to optimize Lignum et Radix Naucleae water or ethanol or ethanol water solution extract, the active stronger extract of exploitation.
Three of object is to disclose the preparation method of said extracted thing.
Technical scheme of the present invention:
One, the preparation method of Lignum et Radix Naucleae extract of the present invention, comprises the following steps:
1. get Lignum et Radix Naucleae medical material, water or ethanol or ethanol water extract, and reclaim solvent and obtain extractum, make Lignum et Radix Naucleae water extract or ethanol extraction or ethanol water solution extract.
2. above-mentioned extractum thin up, then with petroleum ether or ethyl acetate or n-butanol extraction, gets that extract is concentrated makes respectively corresponding Lignum et Radix Naucleae extract.
Described Lignum et Radix Naucleae medical material weight with extract solvent volume than preferred 1:5～15.
Advise with isopyknic extractant, extracting after described extractum thin up.
Lignum et Radix Naucleae 70% ethanol extraction of preferably being prepared by following methods: get Lignum et Radix Naucleae medical material, in medical material weight and solvent volume, than the ratio for 1:10, with 70% ethanol water, extract twice, reclaim solvent and obtain extractum, after extractum thin up, with equal-volume ethyl acetate extraction three times, combining extraction liquid concentrates to obtain described Lignum et Radix Naucleae extract.
Two, using above-mentioned Lignum et Radix Naucleae extract as active component, be applied to the preparation of methicillin-resistant staphylococcus aureus resistance medicine.
The present invention, by Lignum et Radix Naucleae being extracted with separated, screens the pharmacological action of each extract methicillin-resistant staphylococcus aureus resistance (MRSA), investigates the effect of the anti-MRSA of Lignum et Radix Naucleae.Experimental result shows: Lignum et Radix Naucleae water, ethanol, ethanol water solution extract and corresponding ethyl acetate, n-butyl alcohol and water layer extract all have stronger anti-MRSA effect, wherein the acetic acid ethyl ester extract activity of Lignum et Radix Naucleae 70% ethanol extraction is the strongest, is obviously better than extract prepared by additive method.
Above-mentioned Lignum et Radix Naucleae extract of the present invention can be used as the active component of anti-microbial type pharmaceutical preparation, makes tablet, capsule, slow releasing agent, injection, suppository, compound preparation etc.
The present invention has following beneficial effect:
The new purposes of having expanded medical material Lignum et Radix Naucleae with the present invention, the product being prepared by the inventive method has the effect of methicillin-resistant staphylococcus aureus resistance (MRSA), can be used as natural antibacterial, is applied to the industries such as food, medicine; Realize the comprehensive utilization of Lignum et Radix Naucleae plant simultaneously and improve its added value.The alleged medicine of the present invention comprises that people uses medicine and other medicine for animal.
Preparation technology of the present invention is simple, and production cost is low, can adapt to large-scale production.
The specific embodiment
By specific embodiment, further illustrate technical scheme of the present invention below.
The present invention screens the pharmacological action of the methicillin-resistant staphylococcus aureus resistance of Lignum et Radix Naucleae extract by following method.
With micro-broth dilution method Lignum et Radix Naucleae extract to the minimum inhibitory concentration of methicillin-resistant staphylococcus aureus (MIC) and minimum bactericidal concentration (MBC).
Bacterial strain: methicillin-resistant staphylococcus aureus (MRSA is numbered M1-M30) clinical separation strain 30 strains; Staphylococcus aureus reference culture (Sa, ATCC25923) provides ，Jing No.1 Hospital Affiliated to Zhongshan Univ. clinical microbiology laboratory technique chamber to detect its drug resistance of confirmation by No.1 Hospital Affiliated to Zhongshan Univ.'s laboratory medicine portion clinical microbiology laboratory technique chamber.
MH broth bouillon: MH meat soup dry powder (Britain OXOID LTD.) 2.1g, be settled to 100ml, NaOH regulates pH to 7.0, and autoclaving is put 4 ℃ of refrigerators standby.
Method: the micro-broth dilution method operation of recommending with reference to the U.S. state-run clinical trial standardization committee (NCCLS).
Embodiment 1
Preparation Lignum et Radix Naucleae water extract and extract thereof
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is provided by Hainan golden light pharmaceutcal corporation, Ltd.Lignum et Radix Naucleae 3000g, water 45000ml decocts 2 times, each 2 hours, filters; Merging filtrate, concentrates to obtain extractum (water extract) 296.5g.Get extractum 197.7g, after water 2000ml suspendible, with each 2000ml, use respectively petroleum ether, ethyl acetate, n-butanol extraction, every kind of solvent extraction three times, merge respectively each extract, concentrate to obtain ligroin extraction 0.9g, ethyl acetate extract 20.3g, n-butanol extract 74.1g, water layer extract 102.4g.
Embodiment 2
The screening of the pharmacological action of Lignum et Radix Naucleae water extract and extract methicillin-resistant staphylococcus aureus resistance (MRSA) thereof
1 experimental technique
1) mensuration of minimum inhibitory concentration (MIC):
Lignum et Radix Naucleae water extract and extract thereof, erythromycin, ceftriaxone sodium, levofloxacin, vancomycin carry out respectively a series of doubling dilutions in MH broth bouillon, and regulating inoculation bacterium is 1.0 * 10
6cFU/ml.Cultivate 24 hours for 35 ℃, the concentration of the minimum antibacterials that occur without precipitation is its minimum inhibitory concentration (MIC).
2) mensuration of minimum bactericidal concentration (MBC):
From 1) pipette 50 μ l to blood plate the hole of asepsis growth, evenly coating, cultivate 24 hours for 35 ℃, colony counting, making the concentration of initial experiment viable count minimizing 99.9% or above needed minimum antibacterials is its minimum bactericidal concentration (MBC).
By measuring minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of medicine, and data are added up and drawn MIC
50, MIC
90, MBC
50, MBC
90, evaluate the effect of medicine methicillin-resistant staphylococcus aureus resistance.
2 experimental results
Lignum et Radix Naucleae water extract and extract thereof, four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) the results are shown in Table 1 to the extracorporeal bacteria inhibitor test of methicillin-resistant staphylococcus aureus.
Lignum et Radix Naucleae water extract and extract thereof to the extracorporeal disinfecting result of the test of methicillin-resistant staphylococcus aureus in Table 2.
Through statistical analysis, Lignum et Radix Naucleae water extract and extract thereof the MIC to the In Vitro Bacteriostasis of methicillin-resistant staphylococcus aureus and bactericidal assay
50, MIC
90, MBC
50, MBC
90in Table 3.
Through statistical analysis, the MIC of four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) to the bacteriostatic test of methicillin-resistant staphylococcus aureus
50, MIC
90in Table 4:
Table 1 Lignum et Radix Naucleae water extract and extract thereof, the four kinds of antibiotic extracorporeal bacteria inhibitor test result to MRSA
Table 2 Lignum et Radix Naucleae water extract and extract thereof the extracorporeal disinfecting result of the test to MRSA
The statistical result to the In Vitro Bacteriostasis of MRSA and bactericidal assay of table 3 Lignum et Radix Naucleae water extract and extract thereof
The statistical result of four kinds of antibiotic of table 4 to the extracorporeal bacteria inhibitor test of MRSA
Experimental result shows, Lignum et Radix Naucleae water extract and acetic acid ethyl ester extract thereof, n-butyl alcohol extract, water layer have methicillin-resistant staphylococcus aureus resistance (MRSA) effect.Wherein the activity of the anti-MRSA of acetic acid ethyl ester extract of water extract is the strongest.
Embodiment 3
Preparation Lignum et Radix Naucleae 10% ethanol extraction and extract thereof
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is provided by Hainan golden light pharmaceutcal corporation, Ltd.Lignum et Radix Naucleae 3000g, by 10% ethanol 15000ml reflux, extract, 2 times, each 2 hours, filters; Merging filtrate, concentrates to obtain extractum (10% ethanol extraction) 288.3g.Get extractum 192.2g, use respectively petroleum ether, ethyl acetate, n-butanol extraction, every kind of solvent extraction three times, each 2000ml, merge respectively each extract, concentrate to obtain petroleum ether extract 1.0g, acetic acid ethyl ester extract 20.2g, n-butyl alcohol extract 72.7g, water layer extract 98.3g.
Embodiment 4
The screening of the pharmacological action of Lignum et Radix Naucleae 10% ethanol extraction and extract methicillin-resistant staphylococcus aureus resistance (MRSA) thereof
1 experimental technique
3) mensuration of minimum inhibitory concentration (MIC):
Lignum et Radix Naucleae 10% ethanol extraction and extract thereof, erythromycin, ceftriaxone sodium, levofloxacin, vancomycin carry out respectively a series of doubling dilutions in MH broth bouillon, and regulating inoculation bacterium is 1.0 * 10
6cFU/ml.Cultivate 24 hours for 35 ℃, the concentration of the minimum antibacterials that occur without precipitation is its minimum inhibitory concentration (MIC).
4) mensuration of minimum bactericidal concentration (MBC):
From 1) pipette 50 μ l to blood plate the hole of asepsis growth, evenly coating, cultivate 24 hours for 35 ℃, colony counting, making the concentration of initial experiment viable count minimizing 99.9% or above needed minimum antibacterials is its minimum bactericidal concentration (MBC).
By measuring minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of medicine, and data are added up and drawn MIC
50, MIC
90, MBC
50, MBC
90, evaluate the effect of medicine methicillin-resistant staphylococcus aureus resistance.
2 experimental results
Lignum et Radix Naucleae 10% ethanol extraction and extract thereof, four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) the results are shown in Table 5 to the extracorporeal bacteria inhibitor test of methicillin-resistant staphylococcus aureus.
Lignum et Radix Naucleae 10% ethanol extraction and extract thereof to the extracorporeal disinfecting result of the test of methicillin-resistant staphylococcus aureus in Table 6.
Through statistical analysis, Lignum et Radix Naucleae 10% ethanol extraction and the MIC of extract to the In Vitro Bacteriostasis of methicillin-resistant staphylococcus aureus and bactericidal assay thereof
50, MIC
90, MBC
50, MBC
90in Table 7.
Through statistical analysis, the MIC of four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) to the bacteriostatic test of methicillin-resistant staphylococcus aureus
50, MIC
90in Table 8:
Table 5 Lignum et Radix Naucleae 10% ethanol extraction and extract thereof, the four kinds of antibiotic extracorporeal bacteria inhibitor test result to MRSA
Table 6 Lignum et Radix Naucleae 10% ethanol extraction and extract thereof the extracorporeal disinfecting result of the test to MRSA
Table 7 Lignum et Radix Naucleae 10% ethanol extraction and the statistical result of extract to the In Vitro Bacteriostasis of MRSA and bactericidal assay thereof
The statistical result of four kinds of antibiotic of table 8 to the extracorporeal bacteria inhibitor test of MRSA
Experimental result shows, Lignum et Radix Naucleae 10% ethanol extraction and acetic acid ethyl ester extract thereof, n-butyl alcohol extract, water layer have methicillin-resistant staphylococcus aureus resistance (MRSA) effect.Wherein the activity of the anti-MRSA of acetic acid ethyl ester extract of 10% ethanol extraction is the strongest.
Embodiment 5
Preparation Lignum et Radix Naucleae 40% ethanol extraction and extract thereof
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is provided by Hainan golden light pharmaceutcal corporation, Ltd.Lignum et Radix Naucleae 3000g, by 40% ethanol 30000ml reflux, extract, 2 times, each 2 hours, filters; Merging filtrate, concentrates to obtain extractum (40% ethanol extraction) 279.1g.Get extractum 186.1g, use respectively petroleum ether, ethyl acetate, n-butanol extraction, every kind of solvent extraction three times, each 2000ml, merge respectively each extract, concentrate to obtain petroleum ether extract 1.1g, acetic acid ethyl ester extract 25.4g, n-butyl alcohol extract 63.9g, water layer extract 95.7g.
Embodiment 6
The screening of the pharmacological action of Lignum et Radix Naucleae 40% ethanol extraction and extract methicillin-resistant staphylococcus aureus resistance (MRSA) thereof
1 experimental technique
5) mensuration of minimum inhibitory concentration (MIC):
Lignum et Radix Naucleae 40% ethanol extraction and extract thereof, erythromycin, ceftriaxone sodium, levofloxacin, vancomycin carry out respectively a series of doubling dilutions in MH broth bouillon, and regulating inoculation bacterium is 1.0 * 10
6cFU/ml.Cultivate 24 hours for 35 ℃, the concentration of the minimum antibacterials that occur without precipitation is its minimum inhibitory concentration (MIC).
6) mensuration of minimum bactericidal concentration (MBC):
From 1) pipette 50 μ l to blood plate the hole of asepsis growth, evenly coating, cultivate 24 hours for 35 ℃, colony counting, making the concentration of initial experiment viable count minimizing 99.9% or above needed minimum antibacterials is its minimum bactericidal concentration (MBC).
By measuring minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of medicine, and data are added up and drawn MIC
50, MIC
90, MBC
50, MBC
90, evaluate the effect of medicine methicillin-resistant staphylococcus aureus resistance.
2 experimental results
Lignum et Radix Naucleae 40% ethanol extraction and extract thereof, four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) the results are shown in Table 9 to the extracorporeal bacteria inhibitor test of methicillin-resistant staphylococcus aureus.
Lignum et Radix Naucleae 40% ethanol extraction and extract thereof to the extracorporeal disinfecting result of the test of methicillin-resistant staphylococcus aureus in Table 10.
Through statistical analysis, Lignum et Radix Naucleae 40% ethanol extraction and the MIC of extract to the In Vitro Bacteriostasis of methicillin-resistant staphylococcus aureus and bactericidal assay thereof
50, MIC
90, MBC
50, MBC
90in Table 11.
Through statistical analysis, the MIC of four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) to the bacteriostatic test of methicillin-resistant staphylococcus aureus
50, MIC
90in Table 12:
Table 9 Lignum et Radix Naucleae 40% ethanol extraction and extract thereof, the four kinds of antibiotic extracorporeal bacteria inhibitor test result to MRSA
Table 10 Lignum et Radix Naucleae 40% ethanol extraction and extract thereof the extracorporeal disinfecting result of the test to MRSA
Table 11 Lignum et Radix Naucleae 40% ethanol extraction and the statistical result of extract to the In Vitro Bacteriostasis of MRSA and bactericidal assay thereof
The statistical result of four kinds of antibiotic of table 12 to the extracorporeal bacteria inhibitor test of MRSA
Experimental result shows, Lignum et Radix Naucleae 40% ethanol extraction and acetic acid ethyl ester extract thereof, n-butyl alcohol extract, water layer have methicillin-resistant staphylococcus aureus resistance (MRSA) effect.Wherein the activity of the anti-MRSA of acetic acid ethyl ester extract of 40% ethanol extraction is the strongest.
Embodiment 7
Preparation Lignum et Radix Naucleae 70% ethanol extraction and extract thereof
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is provided by Hainan golden light pharmaceutcal corporation, Ltd.Lignum et Radix Naucleae 3000g breaks into coarse powder, with 70% ethanol 30000ml, refluxes 2 times, each 2 hours, filters; Merging filtrate, concentrates to obtain extractum (70% ethanol extraction) 276.0g.Get extractum 184.0g, after water 2000ml suspendible, with each 2000ml, use respectively petroleum ether, ethyl acetate, n-butanol extraction, every kind of solvent extraction three times, merge respectively each extract, concentrate to obtain ligroin extraction 2.1g, ethyl acetate extract 28.3g, n-butanol extract 61.9g, water layer extract 91.7g.
Embodiment 8
The screening of the pharmacological action of Lignum et Radix Naucleae 70% extract and extract methicillin-resistant staphylococcus aureus resistance thereof
1 experimental technique
7) mensuration of minimum inhibitory concentration (MIC):
Lignum et Radix Naucleae 70% ethanol extraction and extract thereof, erythromycin, ceftriaxone sodium, levofloxacin, vancomycin carry out respectively a series of doubling dilutions in MH broth bouillon, and regulating inoculation bacterium is 1.0 * 10
6cFU/ml.Cultivate 24 hours for 35 ℃, the concentration of the minimum antibacterials that occur without precipitation is its minimum inhibitory concentration (MIC).
8) mensuration of minimum bactericidal concentration (MBC):
From 1) pipette 50 μ l to blood plate the hole of asepsis growth, evenly coating, cultivate 24 hours for 35 ℃, colony counting, making the concentration of initial experiment viable count minimizing 99.9% or above needed minimum antibacterials is its minimum bactericidal concentration (MBC).
By measuring minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of medicine, and data are added up and drawn MIC
50, MIC
90, MBC
50, MBC
90, evaluate the effect of medicine methicillin-resistant staphylococcus aureus resistance.
2 experimental results
Lignum et Radix Naucleae 70% ethanol extraction and extract thereof, four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) the results are shown in Table 13 to the extracorporeal bacteria inhibitor test of methicillin-resistant staphylococcus aureus.
Lignum et Radix Naucleae 70% ethanol extraction and extract thereof to the extracorporeal disinfecting result of the test of methicillin-resistant staphylococcus aureus in Table 14.
Through statistical analysis, Lignum et Radix Naucleae 70% ethanol extraction and the MIC of extract to the In Vitro Bacteriostasis of methicillin-resistant staphylococcus aureus and bactericidal assay thereof
50, MIC
90, MBC
50, MBC
90in Table 15.
Through statistical analysis, the MIC of four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) to the bacteriostatic test of methicillin-resistant staphylococcus aureus
50, MIC
90in Table 16:
Table 13 Lignum et Radix Naucleae 70% ethanol extraction and extract thereof, the four kinds of antibiotic extracorporeal bacteria inhibitor test result to MRSA
Table 14 Lignum et Radix Naucleae 70% ethanol extraction and extract thereof the extracorporeal disinfecting result of the test to MRSA
Table 15 Lignum et Radix Naucleae 70% ethanol extraction and the statistical result of extract to the In Vitro Bacteriostasis of MRSA and bactericidal assay thereof
The statistical result of four kinds of antibiotic of table 16 to the extracorporeal bacteria inhibitor test of MRSA
Experimental result shows, Lignum et Radix Naucleae 70% ethanol extraction and acetic acid ethyl ester extract thereof, n-butyl alcohol extract, water layer have stronger methicillin-resistant staphylococcus aureus resistance (MRSA) effect.Wherein the activity of the anti-MRSA of the acetic acid ethyl ester extract of 70% ethanol extraction (being extract of the present invention) is the strongest.
Embodiment 9
Preparation Lignum et Radix Naucleae 95% ethanol extraction and extract thereof
Lignum et Radix Naucleae (Nauclea officinalis Pierre ex Pitard) is provided by Hainan golden light pharmaceutcal corporation, Ltd.Lignum et Radix Naucleae 3000g, by 95% ethanol 30000ml reflux, extract, 2 times, each 2 hours, filters; Merging filtrate, concentrates to obtain extractum (95% ethanol extraction) 231.8g.Get extractum 154.5g, use respectively petroleum ether, ethyl acetate, n-butanol extraction, every kind of solvent extraction three times, each 2000ml, merge respectively each extract, concentrate to obtain petroleum ether extract 3.2g, acetic acid ethyl ester extract 30.0g, n-butyl alcohol extract 42.8g, water layer extract 78.0g.
Embodiment 10
The screening of the pharmacological action of Lignum et Radix Naucleae 95% ethanol extraction and extract methicillin-resistant staphylococcus aureus resistance (MRSA) thereof
1 experimental technique
9) mensuration of minimum inhibitory concentration (MIC):
Lignum et Radix Naucleae 95% ethanol extraction and extract thereof, erythromycin, ceftriaxone sodium, levofloxacin, vancomycin carry out respectively a series of doubling dilutions in MH broth bouillon, and regulating inoculation bacterium is 1.0 * 10
6cFU/ml.Cultivate 24 hours for 35 ℃, the concentration of the minimum antibacterials that occur without precipitation is its minimum inhibitory concentration (MIC).
10) mensuration of minimum bactericidal concentration (MBC):
From 1) pipette 50 μ l to blood plate the hole of asepsis growth, evenly coating, cultivate 24 hours for 35 ℃, colony counting, making the concentration of initial experiment viable count minimizing 99.9% or above needed minimum antibacterials is its minimum bactericidal concentration (MBC).
By measuring minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of medicine, and data are added up and drawn MIC
50, MIC
90, MBC
50, MBC
90, evaluate the effect of medicine methicillin-resistant staphylococcus aureus resistance.
2 experimental results
Lignum et Radix Naucleae 95% ethanol extraction and extract thereof, four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) the results are shown in Table 17 to the extracorporeal bacteria inhibitor test of methicillin-resistant staphylococcus aureus.
Lignum et Radix Naucleae 95% ethanol extraction and extract thereof to the extracorporeal disinfecting result of the test of methicillin-resistant staphylococcus aureus in Table 18.
Through statistical analysis, Lignum et Radix Naucleae 95% ethanol extraction and the MIC of extract to the In Vitro Bacteriostasis of methicillin-resistant staphylococcus aureus and bactericidal assay thereof
50, MIC
90, MBC
50, MBC
90in Table 19.
Through statistical analysis, the MIC of four kinds of antibiotic (erythromycin, ceftriaxone sodium, levofloxacin, vancomycin) to the bacteriostatic test of methicillin-resistant staphylococcus aureus
50, MIC
90in Table 20:
Table 17 Lignum et Radix Naucleae 95% ethanol extraction and extract thereof, the four kinds of antibiotic extracorporeal bacteria inhibitor test result to MRSA
Table 18 Lignum et Radix Naucleae 95% ethanol extraction and extract thereof the extracorporeal disinfecting result of the test to MRSA
Table 19 Lignum et Radix Naucleae 95% ethanol extraction and the statistical result of extract to the In Vitro Bacteriostasis of MRSA and bactericidal assay thereof
The statistical result of four kinds of antibiotic of table 20 to the extracorporeal bacteria inhibitor test of MRSA
Experimental result shows, Lignum et Radix Naucleae 95% ethanol extraction and acetic acid ethyl ester extract thereof, n-butyl alcohol extract, water layer have methicillin-resistant staphylococcus aureus resistance (MRSA) effect.Wherein the activity of the anti-MRSA of acetic acid ethyl ester extract of 95% ethanol extraction is the strongest.
Claims (6)
1. a Lignum et Radix Naucleae extract, it is characterized in that this extract prepared by following methods: get Lignum et Radix Naucleae medical material, by the extraction with aqueous solution of extracting aqueous solvent or ethanol or ethanol, reclaim and extract solvent extractum processed, extractum thin up, use respectively extractant petroleum ether or ethyl acetate or n-butanol extraction, extract concentrates to obtain Lignum et Radix Naucleae extract again.
2. Lignum et Radix Naucleae extract as claimed in claim 1, is characterized in that described Lignum et Radix Naucleae medical material weight and extracts solvent volume than being 1:5～15.
3. Lignum et Radix Naucleae extract as claimed in claim 1, is characterized in that with isopyknic extractant, extracting after described extractum thin up.
4. Lignum et Radix Naucleae extract as claimed in claim 1, it is characterized in that this extract prepared by following methods: get Lignum et Radix Naucleae medical material, in medical material weight and solvent volume, than the ratio for 1:10, with 70% ethanol water, extract twice, reclaim solvent and obtain extractum, after extractum thin up, with equal-volume ethyl acetate extraction three times, combining extraction liquid concentrates to obtain described Lignum et Radix Naucleae extract.
5. the application of Lignum et Radix Naucleae extract in preparing methicillin-resistant staphylococcus aureus resistance class medicine as described in claim 1 to 4.
6. the application of Lignum et Radix Naucleae extract in preparing food antibacterial as described in claim 1 to 4.
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| CN101293038A (en) * | 2007-04-24 | 2008-10-29 | 江苏康缘药业股份有限公司 | Gallbladder leaves extract, preparing method, its preparation and uses |
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| CN104740171A (en) * | 2015-04-20 | 2015-07-01 | 海南黎药堂生物科技开发有限公司 | Traditional Chinese medicine composition, preparation as well as preparation method and use of traditional Chinese medicine composition |
| CN104740171B (en) * | 2015-04-20 | 2019-01-01 | 海南黎药堂生物科技开发有限公司 | A kind of Chinese medicine composition, preparation and preparation method, purposes |
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