CN110302219A - Cortex Pseudolaricis extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug - Google Patents
Cortex Pseudolaricis extract is preparing the application in overriding resistance Acinetobacter bauamnnii drug Download PDFInfo
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Abstract
The invention discloses Cortex Pseudolaricis extracts to prepare the application in overriding resistance Acinetobacter bauamnnii drug.The present invention treats the nematode for having infected drug resistance Acinetobacter bauamnnii using the extract of Golden Larch Bark, show that the extract of Golden Larch Bark has good internal bactericidal effect to drug resistance Acinetobacter bauamnnii, this provides good early-stage study basis to research and develop novel overriding resistance Acinetobacter bauamnnii drug, brings dawn without the available predicament of medicine to solve clinically drug resistance Acinetobacter bauamnnii infection.Using extract the preventing and treating to drug resistance Acinetobacter bauamnnii of Golden Larch Bark, method is quick, cheap, and it is convenient to operate.
Description
Technical field
The present invention relates to field of biotechnology, more particularly, to Cortex Pseudolaricis extract to prepare overriding resistance Bao Man motionless
Application in bacillus drug.
Background technique
Acinetobacter bauamnnii is one of the most common type non-fermented Gram-negative bacteria in acinetobacter, belongs to opportunistic
Bacterium is classified as the superbacteria for needing override to solve by the World Health Organization.The bacterium can be faced in hospital environment with long-term surviving
The distribution of bed department is mainly most with ICU, is secondly Respiratory Medicine.Gerontal patient, Abwehrkraft des Koepers be weak and the patient of critical illness,
And using various store periods and be used for a long time broad-spectrum antibiotic therapy patient be susceptible object.Acinetobacter bauamnnii tool
There is the ability for being quickly obtained and propagating drug resistance, drug resistance Acinetobacter bauamnnii shows worldwide fashion trend, it has also become China
One of most important pathogen of nosocomial infection.It is shown according to CHINET bacterial drug resistance monitoring network data in 2018, China 35
Teaching hospital, family Acinetobacter bauamnnii, which accounts for about, is clinically separated the 14.7% of gram-negative bacteria, is only second to escherichia coli and pneumonia gram
The primary bacterium of thunder.And in terms of to clinical common antibiotics drug resistance, Acinetobacter bauamnnii is to penicillins, cephalosporins, carbon mould
Alkenes, the beta-lactam class antibiotic/compound formulation of beta lactamase restrainer, fluoroquinolones, the resistant rate of aminoglycoside
Reach 45.7-80.4%, only there is lower resistant rate to tigecycline and polymyxins.But polymyxins has biggish kidney
Toxicity and constrain its clinical application, and tigecycline is expensive, and there are some defects for its distribution and metabolism, it would be highly desirable to which development is new
Type overriding resistance Acinetobacter bauamnnii drug.
Heat-clearing and detoxifying herb has accumulated experience abundant in terms of the treatment of infectious diseases.Early in Han dynasty treatise on Febrile Diseases
Just summarize many effective prescriptions, such as Xiexin Tang, gegen qinlian decoction, baitouweng decoction.Think now with many scholars
Treatise on Febrile Diseases as classical works it is practical be exactly an infectious diseases monograph, Qing Dynasty's " pestilence opinion ", " Wen Bing Tiao Bian " etc. write
The appearance of work, so that the Couple herbs of infectious diseases have obtained further enriching, it is anti-infective for further investigation traditional Chinese medicine
Treatment provides precious resources.Domestic and foreign scholars have done a lot of work to Chinese herbal antibacterial Effect study at present, test and face
Bed research all obtains certain achievement.However, these researchs are mostly antibacterial activity in vitro research, to high toxicity, pharmacokinetics
The compound of matter difference and In vitro-in vivo correlation difference lacks effective elimination method, therefore fails to obtain reliable result of study always.
Golden Larch Bark, Chinese medicine name.For the drying of pinaceae plant golden larch Pseudolarix amabilis (Nelson) Rehd.
Root skin or near root bark.It is distributed in the ground such as Jiangsu, Zhejiang, Anhui, Fujian, Jiangxi, Hunan, Hubei, Sichuan.With desinsection, treat
Tinea, it is antipruritic the effect of.Be usually used in scabies itch (as " 3 kinds of Chinese medical extracts to the external Combination susceptibility testing of dermatophyte,
2012, Chen Zhangbao ").It there are no its report to drug-fast bacteria effect at present.
Summary of the invention
The present invention is directed to overcome in the prior art not yet discovery insufficient, research for the effective Chinese medical extract of drug-fast bacteria
It was found that a kind of new internal overriding resistance Acinetobacter bauamnnii drug, i.e. Cortex Pseudolaricis extract.
The first purpose of the invention is to provide Golden Larch Barks to prepare the application in overriding resistance Acinetobacter bauamnnii drug.
The first purpose of the invention is to provide Cortex Pseudolaricis extracts in preparing overriding resistance Acinetobacter bauamnnii drug
Using.
To achieve the goals above, the present invention is achieved by the following technical programs:
Inventor establishes drug resistance Acinetobacter bauamnnii-Caenorhabditis elegans infection model, is extracted using it to Golden Larch Bark
The activity in vivo of the overriding resistance Acinetobacter bauamnnii of object is detected, as the result is shown its can significantly improve Caenorhabditis elegans at
Motility rate illustrates that it has good biological control effect to overriding resistance Acinetobacter bauamnnii, is conducive to Golden Larch Bark and solves superbacteria
Have accumulated good theoretical basis and practical experience.
Therefore claimed Golden Larch Bark is preparing the application in anti-Acinetobacter bauamnnii drug.
The present invention is also claimed Cortex Pseudolaricis extract in anti-Acinetobacter bauamnnii or prepares anti-Acinetobacter bauamnnii drug
In application.
Wherein, the Acinetobacter bauamnnii is drug resistance Acinetobacter bauamnnii.
The anti-Acinetobacter bauamnnii drug is the drug of internal anti-Acinetobacter bauamnnii.
The drug resistance refers to penicillins, cephalosporins, Carbapenems, beta-lactam class antibiotic/beta-lactam
Compound formulation, fluoroquinolones and/or the aminoglycoside medicaments of enzyme inhibitor have drug resistance.
Furthermore it is preferred that the Cortex Pseudolaricis extract is.
It is highly preferred that the preparation method of the water extract of the Golden Larch Bark the following steps are included:
S1. Golden Larch Bark is mixed with water according to mass ratio 1:5~20,90~100 DEG C of 1~2h of heating and refluxing extraction, solid-liquid
Separation；
S2. the obtained solid of step S1 is mixed with water according to mass ratio 1:5~20,90~100 DEG C of heating and refluxing extractions 1
~2h is separated by solid-liquid separation；
S3. merge the liquid that step S1 is obtained and the liquid that step S2 is obtained, removal solvent obtains drug extract.
It is highly preferred that the mass ratio of Golden Larch Bark and water is 1:12 in step S1.
It is highly preferred that extraction conditions are 100 DEG C of extraction 1.5h in step S1 and S2.
It is highly preferred that the mass ratio of solid and water is 1:8 in step S2.
Compared with prior art, the invention has the following beneficial effects:
The invention discloses Cortex Pseudolaricis extracts to prepare the application in overriding resistance Acinetobacter bauamnnii drug.Benefit of the invention
The nematode for having infected drug resistance Acinetobacter bauamnnii is treated with the extract of Golden Larch Bark, shows the extract of Golden Larch Bark to resistance to
Medicine Acinetobacter bauamnnii has good bactericidal effect, this is provided well to research and develop novel overriding resistance Acinetobacter bauamnnii drug
Early-stage study basis, for solve clinically drug resistance Acinetobacter bauamnnii infection bring dawn without the available predicament of medicine.It utilizes
The extract of Golden Larch Bark is prevented and treated drug resistance Acinetobacter bauamnnii, and method is quick, cheap, and it is convenient to operate.
Specific embodiment
The present invention is made below with reference to specification and specific embodiment and further being elaborated, the embodiment is only used
In explaining the present invention, it is not intended to limit the scope of the present invention.Test method as used in the following examples is such as without special theory
It is bright, it is conventional method；Used material, reagent etc., unless otherwise specified, for the reagent and material commercially obtained
Material.
The preparation of 1 Cortex Pseudolaricis extract of embodiment and its detection of bacteriostatic activity
One, experimental method
1, the preparation of Cortex Pseudolaricis extract
Three parts of Golden Larch Barks are weighed, every part of 2g is respectively placed in 3 100mL round-bottomed flasks, by the material ratio of 1:12, respectively
The different solvent 24mL of water, 50% ethyl alcohol, three kinds of 95% ethyl alcohol is added；The flask of three different extracting solutions is respectively set different
Extracting temperature is 100 DEG C, 90 DEG C, 85 DEG C, by medical filtration after heating and refluxing extraction 1.5h；Distinguish again by the material ratio of 1:8
Corresponding extracting solution is added, 100 DEG C, 90 DEG C, 85 DEG C of extraction 1.5h, filter and merge whole medical fluids, be evaporated off under reduced pressure whole respectively
Solvent obtains drug extract, is weighed and placed in 1.5mL centrifuge tube, places in -20 DEG C of refrigerators stand-by.Three kinds of native chaste trees are prepared
Skin extract (water extract of Golden Larch Bark, 50% ethanol extract of Golden Larch Bark, Golden Larch Bark 95% ethanol extract).
2, drug resistance Acinetobacter bauamnnii-Caenorhabditis elegans infection model
It is using 20% brain heart infusion (BHI) fluid nutrient medium containing 10 μm of ol iron ions that synchronized nematode is raw from nematode
It rinsing on long culture medium into 15mL centrifuge tube, 800rpm revolving speed is centrifuged 1min, discards supernatant liquid, and add culture medium to 2mL or so,
Being placed in temperature is 25 DEG C, spare in the insulating box that humidity is 85%；20%BHI is poured into 3mL opacity tube, from Luria-
It is sufficiently mixed in opacity tube with sterile swab stick picking drug resistance Acinetobacter bauamnnii single bacterium colony on Bertan (LB) agar plate
It is even, bacteria concentration is adjusted to 0.5 maxwell unit (about 1 × 10 using than turbid instrument8CFU/mL 250 μ L), then with liquid-transfering gun are sucked out extremely
In new 5mL centrifuge tube, 2250 μ L of 20%BHI fluid nutrient medium is added, bacterium solution is diluted 10 times to 1 × 107CFU/mL；With
The bacterium solution that the liquid-transfering gun of 1000 μ L draws after 2mL dilution is added in nematode pipe, makes final concentration of the 5 of drug resistance Acinetobacter bauamnnii
×106CFU/mL, being put into temperature later is 25 DEG C, is cultivated in the constant temperature and humidity incubator that humidity is 85%；It is incubated for infection 6h altogether
Afterwards, it is cleaned at least 3 times with M9 fluid nutrient medium, 180 μ L of draw solution is assigned in 96 orifice plates, and 15~20, every hole is advisable.System
It is standby to obtain drug resistance Acinetobacter bauamnnii-Caenorhabditis elegans infection model.
3, Cortex Pseudolaricis extract antibacterial activity in vivo detects
The drug resistance Acinetobacter bauamnnii being prepared using previous step-Caenorhabditis elegans infection model is carried out Golden Larch Bark and mentioned
Object antibacterial activity in vivo is taken to detect.
Using the polymyxin B of 20 μ g/mL as positive controls, using 10% DMSO as negative control group, with difference
Golden Larch Bark (water, 50% ethyl alcohol, 95% second of concentration (10000 μ g/mL, 1000 μ g/mL, 100 μ g/mL, 10 μ g/mL, 1 μ g/mL)
Alcohol) for extract as experimental group, every group sets 3 groups of parallel controls.
The Cortex Pseudolaricis extract preparation method of various concentration: weighing the extract of 10mg into 1.5mL centrifuge tube, is added
The DMSO of 100 μ L, the BHI culture medium for adding 900 μ L are sufficiently mixed dissolution extract, and 100 μ L are then successively sucked out and are added
900 μ L BHI culture mediums are serially diluted 5 times, and respectively 10000 μ g/mL, 1000 μ g/mL, 100 μ g/mL, 10 μ g/mL, 1 μ are obtained
The extracting solution of g/mL.
20 μ L Cortex Pseudolaricis extracts are separately added into corresponding aperture, make Cortex Pseudolaricis extract with 1000 μ g/mL, 100 μ g/
ML, 10 μ g/mL, 1 μ g/mL, 0.1 μ g/mL gradient final concentration extract intervened；After being incubated for 36h in 25 DEG C of insulating box,
Therapeutic effect of the Chinese medical extract to infection nematode of various concentration is judged by the survival rate of nematode.With negative control and the positive
Control is as control.
Two, experimental result is shown in Table 1.
Table 1:
The results show that Cortex Pseudolaricis extract, including water extract and alcohol extract, there is obvious overriding resistance Bao Man in vivo not
The effect of lever bacterium, wherein the water extract of Golden Larch Bark has the effect of better biological control nematode relative to ethanol extract,
The survival rate of infection nematode can reach 45% or more.
The preparation of 1 forsythia suspense extraction of reference examples and its detection of bacteriostatic activity
One, experimental method
The preferable forsythia suspense extraction of external overriding resistance Acinetobacter bauamnnii activity being had verified that with this field is control,
The preparation and its detection of bacteriostatic activity in vivo of forsythia suspense extraction are carried out according to 1 method of embodiment.
Two, experimental result is shown in Table 2.
Table 2:
The results show that the results show that forsythia suspense extraction influences very little to the survival rate of infection nematode, without internal overriding resistance Bao
The effect of graceful acinetobacter calcoaceticus.
Claims (10)
1. Golden Larch Bark is in anti-Acinetobacter bauamnnii or prepares application in anti-Acinetobacter bauamnnii drug.
2. Cortex Pseudolaricis extract is in anti-Acinetobacter bauamnnii or prepares application in anti-Acinetobacter bauamnnii drug.
3. application according to claim 1 or claim 2, which is characterized in that the anti-Acinetobacter bauamnnii drug is internal anti-Bao Man
The drug of acinetobacter calcoaceticus.
4. application according to claim 1 or claim 2, which is characterized in that the Acinetobacter bauamnnii is drug resistance Acinetobacter bauamnnii.
5. applying according to claim 4, which is characterized in that the drug resistance refers to green to penicillins, cephalosporins, carbon
Mould alkenes, beta-lactam class antibiotic/beta lactamase restrainer compound formulation, fluoroquinolones and/or aminoglycoside medicine
Object has drug resistance.
6. application according to claim 2, which is characterized in that the Cortex Pseudolaricis extract is the water extract of Golden Larch Bark.
7. applying according to claim 6, which is characterized in that the preparation method of the Cortex Pseudolaricis extract includes following step
It is rapid:
S1. Golden Larch Bark is mixed according to mass ratio 1:5~20,90~100 DEG C of 1~2h of heating and refluxing extraction with water, is separated by solid-liquid separation；
S2. the obtained solid of step S1 is mixed with water according to mass ratio 1:5~20,90~100 DEG C of heating and refluxing extractions 1~
2h is separated by solid-liquid separation；
S3. merge the liquid that step S1 is obtained and the liquid that step S2 is obtained, removal solvent obtains drug extract.
8. applying according to claim 7, which is characterized in that in step S1, the mass ratio of Golden Larch Bark and water is 1:12.
9. applying according to claim 7, which is characterized in that extraction conditions are 100 DEG C of extraction 1.5h in step S1 and S2.
10. applying according to claim 7, which is characterized in that in step S2, the mass ratio of solid and water is 1:8.
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