CN110302219B - Application of cortex pseudolaricis extract in preparation of drug-resistant Acinetobacter baumannii drug - Google Patents
Application of cortex pseudolaricis extract in preparation of drug-resistant Acinetobacter baumannii drug Download PDFInfo
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- FPZLLRFZJZRHSY-HJYUBDRYSA-N tigecycline Chemical compound C([C@H]1C2)C3=C(N(C)C)C=C(NC(=O)CNC(C)(C)C)C(O)=C3C(=O)C1=C(O)[C@@]1(O)[C@@H]2[C@H](N(C)C)C(O)=C(C(N)=O)C1=O FPZLLRFZJZRHSY-HJYUBDRYSA-N 0.000 description 3
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/13—Coniferophyta (gymnosperms)
- A61K36/15—Pinaceae (Pine family), e.g. pine or cedar
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
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- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation or decoction
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Abstract
The invention discloses application of a cortex pseudolaricis extract in preparing a drug-resistant Acinetobacter baumannii drug. According to the invention, the extract of the cortex pseudolaricis is used for treating nematodes infected with drug-resistant acinetobacter baumannii, and the extract of the cortex pseudolaricis has a good in-vivo sterilization effect on the drug-resistant acinetobacter baumannii, so that a good early-stage research foundation is provided for developing novel drug-resistant acinetobacter baumannii drugs, and the problem that no drug is available for clinical drug-resistant acinetobacter baumannii infection is solved. The extract of the golden larch bark is utilized to prevent and treat drug-resistant Acinetobacter baumannii, and the method is quick, low in price and convenient to operate.
Description
Technical Field
The invention relates to the technical field of biology, in particular to application of a cortex pseudolaricis extract in preparing a drug-resistant Acinetobacter baumannii drug.
Background
Acinetobacter baumannii is the most common non-fermenting gram-negative bacterium in Acinetobacter, belongs to conditional pathogenic bacteria, and is listed by the world health organization as superbacteria which need to be solved most preferentially. The strain can survive for a long time in a hospital environment, and clinical departments mainly use ICU most, and secondly respiratory medicine. Elderly patients, patients with weak body resistance and critical illness, and patients treated with various invasive procedures and long-term use of broad-spectrum antibiotics are susceptible subjects. Acinetobacter baumannii has the capability of rapidly acquiring and transmitting drug resistance, and the drug-resistant Acinetobacter baumannii presents a worldwide epidemic trend, and becomes one of the most important pathogenic bacteria for infection in China. According to the data of the CHINET bacterial drug resistance monitoring network in 2018, acinetobacter baumannii in 35 education academy of China accounts for about 14.7% of clinically isolated gram-negative bacteria, and is second only to Escherichia coli and Klebsiella pneumoniae. In the aspect of drug resistance to common clinical antibiotics, the drug resistance rate of the acinetobacter baumannii to penicillin, cephalosporin, carbapenem, beta lactam antibiotics/beta lactamase inhibitor composite preparation, fluoroquinolones and aminoglycosides reaches 45.7-80.4 percent, and the acinetobacter baumannii has lower drug resistance rate only to tigecycline and polymyxin. However, polymyxin has great nephrotoxicity, which restricts the clinical application, tigecycline is expensive, and the distribution metabolism of tigecycline has some defects, so that development of novel drug-resistant acinetobacter baumannii drugs is urgent.
The traditional Chinese medicine for clearing heat and detoxicating has accumulated a lot of experience in the treatment of infectious diseases. Many effective prescriptions such as XIE decoction, GEGENQINLIAN decoction, and radix Pulsatillae decoction are summarized in Han dynasty's treatise on typhoid fever'. At present, a plurality of students consider that the typhoid theory is actually a monograph of infectious diseases as classical works, and the works of the epidemic disease theory, the epidemic disease theory and the epidemic disease theory appear in the past, so that the treatment experience of the infectious diseases is further enriched, and precious resources are provided for the intensive study of the anti-infection treatment of traditional Chinese medicines. At present, scholars at home and abroad have made a great deal of work on the research on the antibacterial action of the Chinese herbal medicine, and certain results are achieved in experiments and clinical researches. However, most of these studies are in vitro antibacterial activity studies, and efficient elimination methods are lacking for compounds with high toxicity, poor pharmacokinetic properties and poor in vivo and in vitro correlation, so reliable research results have not been obtained yet.
Cortex pseudolaricis, and Chinese medicine. Is dried root bark or near root bark of Pinaceae plant herba Desmodii Styracifolii Pseudolarix amabilis (Nelson) Rehd. Distributed in Jiangsu, zhejiang, anhui, fujian, jiangxi, hunan, hubei, sichuan, etc. Has effects of killing parasite, treating tinea, and relieving itching. Is commonly used for itch and pruritus (for example, 3 traditional Chinese medicine extracts are used for in vitro combined drug sensitivity test of dermatophytes, 2012, chen Zhangbao). At present, no report on the effect of the strain on drug-resistant bacteria exists.
Disclosure of Invention
The invention aims to overcome the defect that an effective traditional Chinese medicine extract aiming at drug-resistant bacteria is not found in the prior art, and researches and discovers a novel in-vivo drug-resistant Acinetobacter baumannii medicine, namely an extract of golden larch bark.
The first object of the invention is to provide an application of golden larch bark in preparing medicine for resisting drug-resistant acinetobacter baumannii.
The first object of the invention is to provide an application of the cortex pseudolaricis extract in preparing medicine for resisting drug-resistant Acinetobacter baumannii.
In order to achieve the above object, the present invention is realized by the following technical scheme:
the inventor establishes a drug-resistant Acinetobacter baumannii-caenorhabditis elegans infection model, and utilizes the drug-resistant Acinetobacter baumannii to detect the in-vivo activity of the drug-resistant Acinetobacter baumannii of the golden larch bark extract, and the result shows that the drug-resistant Acinetobacter baumannii infection model can remarkably improve the survival rate of the caenorhabditis elegans, so that the drug-resistant Acinetobacter baumannii has a good biological control effect on the drug-resistant Acinetobacter baumannii, and is favorable for solving the accumulation of superbacteria by the golden larch bark.
Therefore, the invention claims the application of the golden larch bark in preparing the medicine for resisting Acinetobacter baumannii.
The invention also claims the application of the cortex pseudolaricis extract in resisting Acinetobacter baumannii or preparing medicines for resisting Acinetobacter baumannii.
Wherein the Acinetobacter baumannii is drug-resistant Acinetobacter baumannii.
The anti-Acinetobacter baumannii drug is an in-vivo anti-Acinetobacter baumannii drug.
The drug resistance refers to drug resistance to penicillin, cephalosporin, carbapenem, beta lactam antibiotics/beta lactamase inhibitor compound preparation, fluoroquinolone and/or aminoglycoside drugs.
In addition, preferably, the golden larch bark extract is.
More preferably, the preparation method of the water extract of cortex pseudolaricis comprises the following steps:
s1, according to the mass ratio of 1: mixing the golden larch bark with water 5-20, heating and refluxing at 90-100 ℃ for 1-2 h, and carrying out solid-liquid separation;
s2, according to the mass ratio of 1: 5-20, mixing the solid obtained in the step S1 with water, heating and refluxing at 90-100 ℃ for 1-2 h, and carrying out solid-liquid separation;
s3, combining the liquid obtained in the step S1 and the liquid obtained in the step S2, and removing the solvent to obtain the medicine extract.
More preferably, in step S1, the mass ratio of cortex pseudolaricis to water is 1:12.
more preferably, the extraction conditions in steps S1 and S2 are 100℃for 1.5h.
More preferably, in step S2, the mass ratio of solids to water is 1:8.
compared with the prior art, the invention has the following beneficial effects:
the invention discloses application of a cortex pseudolaricis extract in preparing a drug-resistant Acinetobacter baumannii drug. According to the invention, the extract of the cortex pseudolaricis is used for treating nematodes infected with drug-resistant acinetobacter baumannii, and the extract of the cortex pseudolaricis has a good bactericidal effect on the drug-resistant acinetobacter baumannii, so that a good early-stage research foundation is provided for developing novel drug-resistant acinetobacter baumannii drugs, and the problem of non-drug-resistant and usable dilemma caused by clinical drug-resistant acinetobacter baumannii infection is solved. The extract of the golden larch bark is utilized to prevent and treat drug-resistant Acinetobacter baumannii, and the method is quick, low in price and convenient to operate.
Detailed Description
The invention is further illustrated in detail in the following description and in the detailed examples, which are provided for the purpose of illustration only and are not intended to limit the scope of the invention. The test methods used in the following examples are conventional methods unless otherwise specified; the materials, reagents and the like used, unless otherwise specified, are those commercially available.
Example 1 preparation of cortex pseudolaricis extract and detection of its antibacterial Activity
1. Experimental method
1. Preparation of cortex pseudolaricis extract
Three parts of golden larch bark are weighed, each 2g is respectively placed in 3 100mL round bottom flasks, and three different solvents of 24mL of water, 50% ethanol and 95% ethanol are respectively added according to the material ratio of 1:12; setting different extraction temperatures of 100 deg.C, 90 deg.C and 85 deg.C in three flasks of different extraction solutions, respectively, heating and reflux extracting for 1.5 hr, and filtering the liquid medicine; and respectively adding corresponding extracting solutions according to the material ratio of 1:8, respectively extracting at 100 ℃ and 90 ℃ and 85 ℃ for 1.5 hours, filtering and merging all the liquid medicine, decompressing and evaporating all the solvent to obtain a medicine extract, weighing and placing in a 1.5mL centrifuge tube, and placing in a refrigerator at-20 ℃ for standby. Three extracts of golden larch bark (water extract of golden larch bark, 50% ethanol extract of golden larch bark, 95% ethanol extract of golden larch bark) were prepared.
2. Drug-resistant acinetobacter baumannii-caenorhabditis elegans infection model
Washing synchronized nematodes from nematode growth medium into 15mL centrifuge tube by using 20% Brain Heart Infusion (BHI) liquid medium containing 10 μmol iron ions, centrifuging at 800rpm for 1min, discarding supernatant, adding medium to about 2mL, and placing in an incubator with temperature of 25deg.C and humidity of 85%; pouring 20% BHI into 3mL turbidimetric tube, picking single colony of drug-resistant Acinetobacter baumannii from Luria-Bertan (LB) agar plate with sterile cotton stick, mixing thoroughly in turbidimetric tube, and adjusting bacterial concentration to 0.5 McO (about 1×10) 8 CFU/mL), then, 250. Mu.L was sucked out into a new 5mL centrifuge tube by a pipette, 2250. Mu.L of 20% BHI liquid medium was added, and the bacterial solution was diluted 10 times to 1X 10 7 CFU/mL; 2mL of diluted bacterial liquid is sucked by a 1000 mu L pipette and added into a nematode tube, so that the final concentration of the drug-resistant Acinetobacter baumannii is 5 multiplied by 10 6 CFU/mL, and then culturing in a constant temperature and humidity incubator with the temperature of 25 ℃ and the humidity of 85%; after 6h of co-incubation of infection, the cells are washed at least 3 times with M9 liquid medium, 180. Mu.L of the aspirated solution is dispensed into 96 well plates, preferably 15 to 20 wells per well. Preparation of drug-resistant Bowman's dropA model of infection with a. Mobilis-caenorhabditis elegans.
3. Antibacterial activity detection in golden larch bark extract
And (3) detecting the in-vivo antibacterial activity of the cortex pseudolaricis extract by using the drug-resistant Acinetobacter baumannii-caenorhabditis elegans infection model prepared in the previous step.
Polymyxin B at 20 μg/mL was used as a positive control group, DMSO at 10% was used as a negative control group, and extracts of golden larch bark (water, 50% ethanol, 95% ethanol) at different concentrations (10000 μg/mL, 1000 μg/mL, 100 μg/mL, 10 μg/mL, 1 μg/mL) were used as experimental groups, each group being provided with 3 parallel controls.
The preparation method of the cortex pseudolaricis extract with different concentrations comprises the following steps: 10mg of the extract was weighed into a 1.5mL centrifuge tube, 100. Mu.L of DMSO was added, 900. Mu.L of BHI medium was added to thoroughly mix and dissolve the extract, then 100. Mu.L of BHI medium was sequentially aspirated and serial dilutions of 900. Mu.L of BHI medium were made 5 times to obtain 10000. Mu.g/mL, 1000. Mu.g/mL, 100. Mu.g/mL, 10. Mu.g/mL, 1. Mu.g/mL of extract, respectively.
Adding 20. Mu.L of cortex pseudolaricis extract into corresponding wells, respectively, and allowing the cortex pseudolaricis extract to interfere with the extract at final concentration of 1000. Mu.g/mL, 100. Mu.g/mL, 10. Mu.g/mL, 1. Mu.g/mL, and 0.1. Mu.g/mL; after incubation for 36h in an incubator at 25 ℃, the treatment effect of the traditional Chinese medicine extracts with different concentrations on the infected nematodes is judged according to the survival rate of the nematodes. Negative and positive controls were used as controls.
2. The experimental results are shown in Table 1.
Table 1:
the results show that the cortex pseudolaricis extract, including the water extract and the alcohol extract, has obvious in-vivo drug-resistant Acinetobacter baumannii effect, wherein the water extract of the cortex pseudolaricis has better biological effect of preventing and treating nematodes relative to the alcohol extract, and the survival rate of the infected nematodes can reach more than 45%.
Comparative example 1 preparation of Forsythia suspensa extract and detection of antibacterial Activity thereof
1. Experimental method
The preparation of the fructus forsythiae extract and the detection of the in-vivo antibacterial activity thereof are carried out according to the method of the embodiment 1 by taking the fructus forsythiae extract with better in-vitro anti-drug-resistant Acinetobacter baumannii activity verified in the field as a control.
2. The experimental results are shown in Table 2.
Table 2:
the results show that the fructus forsythiae extract has little influence on the survival rate of the infected nematodes and has no effect of resisting drug-resistant Acinetobacter baumannii in vivo.
Claims (5)
1. The application of the cortex pseudolaricis extract in preparing the anti-Acinetobacter baumannii medicine is characterized in that the anti-Acinetobacter baumannii medicine is an in-vivo anti-Acinetobacter baumannii medicine;
the golden larch bark extract is an aqueous extract of golden larch bark,
the Acinetobacter baumannii is drug-resistant Acinetobacter baumannii,
the drug resistance refers to drug resistance to penicillin, cephalosporin, carbapenem, beta lactam antibiotics/beta lactamase inhibitor compound preparation, fluoroquinolone and/or aminoglycoside drugs.
2. The use according to claim 1, characterized in that the preparation method of the golden larch bark extract comprises the following steps: s1, according to the mass ratio of 1: mixing the golden larch bark with water 5-20, heating and refluxing at 90-100 ℃ for 1-2 h, and carrying out solid-liquid separation;
s2, according to the mass ratio of 1: 5-20, mixing the solid obtained in the step S1 with water, heating and refluxing at 90-100 ℃ for 1-2 h, and carrying out solid-liquid separation;
s3, combining the liquid obtained in the step S1 and the liquid obtained in the step S2, and removing the solvent to obtain the medicine extract.
3. The use according to claim 2, wherein in step S1, the mass ratio of cortex pseudolaricis to water is 1:12.
4. the use according to claim 2, wherein the extraction conditions in steps S1 and S2 are 100 ℃ for 1.5h.
5. The use according to claim 2, wherein in step S2 the mass ratio of solids to water is 1:8.
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|---|---|---|---|---|
| CN103040804A (en) * | 2012-12-19 | 2013-04-17 | 程新明 | Norwogonin and application thereof to preparation of medicines for treating acinetobacter baumannii infection |
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