CN107648214A - A kind of combined method for improving chloramphenicol effect of drugs - Google Patents
A kind of combined method for improving chloramphenicol effect of drugs Download PDFInfo
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- CN107648214A CN107648214A CN201710928120.1A CN201710928120A CN107648214A CN 107648214 A CN107648214 A CN 107648214A CN 201710928120 A CN201710928120 A CN 201710928120A CN 107648214 A CN107648214 A CN 107648214A
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- chloramphenicol
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
- A61K31/19—Carboxylic acids, e.g. valproic acid
- A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
- A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid, pantothenic acid
- A61K31/198—Alpha-aminoacids, e.g. alanine, edetic acids [EDTA]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/16—Amides, e.g. hydroxamic acids
- A61K31/165—Amides, e.g. hydroxamic acids having aromatic rings, e.g. colchicine, atenolol, progabide
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The present invention relates to a kind of combined method for improving chloramphenicol effect of drugs.Comprise the following steps：A. chloramphenicol is prepared；The preparation of b.N acetylcysteines；The preparation of c.LB fluid nutrient mediums；D. the solid substrate flat board of chloramphenicol is free of；E. the culture of Escherichia coli；F. the dilution of Escherichia coli；G. semi-solid upper panel is fallen without chloramphenicol；H. good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator, bacterium colony is counted.A kind of combined method advantage for improving chloramphenicol effect of drugs of the present invention：The usage amount of chloramphenicol is reduced, reduction amount is 5mg/L chloramphenicol；Eliminate the risk that drug resistance is produced during the chloramphenicol use of low dosage.
Description
Technical field
The present invention relates to a kind of combined method for improving chloramphenicol effect of drugs.
Background technology
The chemical constitution of chloramphenicol contains p-nitrophenyl, propane diols and dichloro acetamide, and its antibacterial activity is mainly with third
Glycol is relevant；For its form to be white to slightly yellow fine acicular or flaky crystal, odorless, taste is extremely bitter, is insoluble in water, is soluble in second
Alcohol, acetone, are slightly soluble in benzene and petroleum ether.Chloramphenicol is trained from streptomyces venezuelae (Streptomyces venezuela) earliest
Separation and Extraction in nutrient solution, belong to broad-spectrum antibiotic class, such as to many gram-positive bacterias and Gram-negative bacteria, bacteroid, Garrick
Ci Shi bodies, Chlamydia etc. have inhibitory action, especially there is good antibacterial to Salmonella, Bacillus influenzae and Bacteroides etc.
Ability, clinically it is mainly used in the treatment of above-mentioned infectious diseases and typhoid fever.The principle that chloramphenicol plays drug effect is chloramphenicol
The 50S subunits of bacterium ribosome are acted on, so as to prevent the synthesis of bacterioprotein.
However, under chloramphenicol coercion various bacteria can to chloramphenicol produce drug resistance, wherein with Escherichia coli,
Shigella dysenteriae, proteus etc. are more common, and typhoid bacillus and staphylococcus are more rare.Bacterial drug resistance means bacterium for anti-
The pharmaceutically-active tolerance of bacterium, once producing, the therapeutic action of medicine is just decreased obviously drug resistance；And when this kind of prolonged application
During antibiotic, the sensitive strain to occupy the majority is constantly killed, and antibody-resistant bacterium instead of sensitive strain, and makes bacterium with regard to amount reproduction
The resistant rate of this kind of medicine is constantly raised.
The drug resistance of bacterium is just found early in nineteen sixty, such as the staphylococcus aureus (MRSA) of methicillin-resistant, with
The green pus bar of the Multiple Classes of Antibiotics such as the enterococcus of vancomycin resistance, resistance to ampicillin, Amoxicillin, Xi Lixin has been found afterwards
The Klebsiella Pneumoniae of 16 kinds of antibiotic such as bacterium, Nai Xilixin, fortum；And the 2010 novel super drug-fast bacterias found in South Asia
NDM-1 (New Delhi metallo-β-lactamase 1, New Delhi metallo- β-lactamase-1) is almost to all β-interior
Amide-type antibiotic all has drug resistance；CDC has also circulated a notice of three and has infected super resistance pathogenic bacteria
Case.
The content of the invention
The problem to be solved in the present invention there is provided a kind of combined method of raising chloramphenicol effect of drugs with environmental protection.
A kind of combined method for improving chloramphenicol effect of drugs of the present invention, it is characterised in that comprise the following steps：
(1) sorting：From
A kind of combined method for improving chloramphenicol effect of drugs, it is characterised in that comprise the following steps：
A. chloramphenicol is prepared：The chloramphenicol mother liquor for being 20mg/mL with absolute ethyl alcohol compound concentration, and with 0.22 μm of filter membrane
Filtration sterilization, it is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of b.N- acetylcysteines：10mg/mL N-acetylcystein is prepared with deionized water and with 0.22 μ
M membrane filtration is degerming, is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of c.LB fluid nutrient mediums：Sodium chloride 5g, peptone 10g, dusty yeast 5g, add water to 1000mL, pH naturally,
Packing is after 1.034 × 105Pa sterilizing 20min, solid medium add w/v 1.2% agar powder thereto；Semisolid training
Support base addition w/v 0.5% agar powder；
D. the solid substrate flat board of chloramphenicol is free of：LB solid mediums after sterilizing fall after it is cooled to 50 DEG C or so
Enter sterile disposable plate, cooling is stand-by；
Bottom solid plate containing chloramphenicol：LB solid mediums after sterilizing after it is cooled to 50 DEG C or so thereto
Chloramphenicol mother liquor is added to the working concentration needed, sterile disposable plate is poured into after mixing, cooling is stand-by；
E. the culture of Escherichia coli：E. coli broth to fresh LB liquid is inoculated with according to the inoculum concentrations of w/v 2% to train
Support and cultivated in base in 37 DEG C of 180rpm to logarithmic phase, it is stand-by；
F. the dilution of Escherichia coli：The Escherichia coli of step e culture are taken to dilute 105It is stand-by again；
G. semi-solid upper panel is fallen without chloramphenicol：50 μ L are taken to dilute 105Escherichia coli again are added in ampere bottle,
The semisolid of 5mL sterilizings is added into ampere bottle again, semi-solid temperature pours into solid plate good in advance at 45 DEG C after shaking up
On；
Semi-solid upper panel is fallen containing chloramphenicol：50 μ L are taken to dilute 105Times Escherichia coli be added in ampere bottle, to
The semisolid of 5mL sterilizings is added in ampere bottle, it is dense to what is needed that semi-solid temperature adds chloramphenicol at 45 DEG C, and in semisolid
Degree, poured into after shaking up on the solid plate of the advance good chloramphenicol containing same concentrations；
H. good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator, bacterium colony is counted.
China is antibiotics production and the big country used, therefore may also turn into the district occurred frequently of bacterial drug resistance, is rationally used
Medicine, Scientific Usage of Drugs are extremely urgent.It is contemplated that improving chloramphenicol medication drug effect and drug safety, it is mould to be related to a kind of chlorine
The drug regimen of element.Composition of medicine, which works well, to be shown to the experiment of Escherichia coli extracorporeal disinfecting, i.e. the chloramphenicol and N- of low dosage
The drug regimen of acetylcysteine.The drug regimen of optimum efficiency is that 1~5mg/L N-acetylcystein+10mg/L chlorine is mould
Element.
A kind of combined method purpose for improving chloramphenicol effect of drugs of the present invention：Chloramphenicol is broad-spectrum antibiotic, in clinic
It is widely used in therapeutic process, but not only dosage is big but also repeated multiple times use can cause bacterial resistance for the chloramphenicol used
The appearance of property.Experiment in vitro experiment shows：15mg/L chloramphenicol can suppress the growth of sensitive E. coli, but low dosage
Chloramphenicol such as 5~10mg/L and lower concentration chloramphenicol can cause being produced to chloramphenicol resistance for part Escherichia coli, from
And cause therapeutic effect bad.The object of the invention is exactly to improve under the chloramphenicol effect of low dosage, reduces Escherichia coli resistance
Property generation, improve therapeutic effect.
A kind of combined method effect for improving chloramphenicol effect of drugs of the present invention：1~5mg/L N-acetylcysteins+
The drug regimen of 10mg/L chloramphenicol can completely inhibit the growth of sensitive E. coli, and not produce drug resistance.
A kind of combined method advantage for improving chloramphenicol effect of drugs of the present invention：The usage amount of chloramphenicol is reduced, is reduced
Measure as 5mg/L chloramphenicol；Eliminate the risk that drug resistance is produced during the chloramphenicol use of low dosage, chloramphenicol containing 10mg/L
The flat board of concentration, the clump count of Escherichia coli is 28 ± 7, in the medicine of 1~5mg/L N-acetylcystein+10mg/L chloramphenicol
The flat board of thing combination, clump count 0.
Embodiment
Below in conjunction with the embodiment of the present invention, the technical scheme in the embodiment of the present invention is clearly and completely described,
Obviously, described embodiment is only part of the embodiment of the present invention, rather than whole embodiments.Based in the present invention
Embodiment, the every other embodiment that those of ordinary skill in the art are obtained under the premise of creative work is not made, all
Belong to the scope of protection of the invention.
Embodiment 1：A kind of combined method for improving chloramphenicol effect of drugs, comprises the following steps：
A. chloramphenicol is prepared：The chloramphenicol mother liquor for being 20mg/mL with absolute ethyl alcohol compound concentration, and with 0.22 μm of filter membrane
Filtration sterilization, it is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of b.N- acetylcysteines：10mg/mL N-acetylcystein is prepared with deionized water and with 0.22 μ
M membrane filtration is degerming, is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of c.LB fluid nutrient mediums：Sodium chloride 5g, peptone 10g, dusty yeast 5g, add water to 1000mL, pH naturally,
Packing is after 1.034 × 105Pa sterilizing 20min, solid medium add w/v 1.2% agar powder thereto；Semisolid training
Support base addition w/v 0.5% agar powder；
D. the solid substrate flat board of chloramphenicol is free of：LB solid mediums after sterilizing fall after it is cooled to 50 DEG C or so
Enter sterile disposable plate, cooling is stand-by；
Bottom solid plate containing chloramphenicol：LB solid mediums after sterilizing after it is cooled to 50 DEG C or so thereto
Chloramphenicol mother liquor is added to the working concentration needed, sterile disposable plate is poured into after mixing, cooling is stand-by；
E. the culture of Escherichia coli：E. coli broth to fresh LB liquid is inoculated with according to the inoculum concentrations of w/v 2% to train
Support and cultivated in base in 37 DEG C of 180rpm to logarithmic phase, it is stand-by；
F. the dilution of Escherichia coli：The Escherichia coli of step e culture are taken to dilute 105It is stand-by again；
G. semi-solid upper panel is fallen without chloramphenicol：50 μ L are taken to dilute 105Escherichia coli again are added in ampere bottle,
The semisolid of 5mL sterilizings is added into ampere bottle again, semi-solid temperature pours into solid plate good in advance at 45 DEG C after shaking up
On；
Semi-solid upper panel is fallen containing chloramphenicol：50 μ L are taken to dilute 105Times Escherichia coli be added in ampere bottle, to
The semisolid of 5mL sterilizings is added in ampere bottle, it is dense to what is needed that semi-solid temperature adds chloramphenicol at 45 DEG C, and in semisolid
Degree, poured into after shaking up on the solid plate of the advance good chloramphenicol containing same concentrations；
H. good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator, bacterium colony is counted.
Embodiment 2：A kind of combined method for improving chloramphenicol effect of drugs, comprises the following steps：
(1) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate is placed in quiescent culture 36h in 37 DEG C of constant incubator (double-layer plate is free of chloramphenicol), counts E. coli clones number,
As a control group.
(2) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate (contains chloramphenicol in double-layer plate and levels concentration is consistent).It is divided into 4 groups, the chloramphenicol concentration that each group of double-layer plate contains
Difference, respectively 5mg/L, 10mg/L, 15mg/L, 20mg/L, good double-layer plate are placed in quiet in 37 DEG C of constant incubator
Put culture 36h and count respectively.
(3) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate (contains N-acetylcystein) in double-layer plate, be divided into 3 groups, and the chloramphenicol concentration that each group of double-layer plate contains is inconsistent,
Respectively 5mg/L, 10mg/L, 15mg/L, good double-layer plate are placed in quiescent culture 36h in 37 DEG C of constant incubator and divided
Do not count.
After above-mentioned different disposal, the clump count of Escherichia coli is as shown in the table：
As shown by data 10mg/L chloramphenicol has 50% lethal effect, the N- mucolyticums of selection to Escherichia coli in table
Acid concentration (5,10,15mg/L) has no significant effect to Escherichia coli Growth.Illustrate concentration list of the N-acetylcystein in experiment
Solely had no significant effect using Escherichia coli.
Embodiment 3：A kind of combined method for improving chloramphenicol effect of drugs, comprises the following steps：
(1) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate is placed in quiescent culture 36h in 37 DEG C of constant incubator (double-layer plate is free of chloramphenicol), counts E. coli clones number,
As a control group.Bacterium colony count results are 59 ± 5.
(2) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, in double-layer plate containing chloramphenicol and levels concentration it is consistent and be 10mg/L, good double-layer plate is placed in 37 DEG C of constant temperature
Quiescent culture 36h and counted in incubator.Bacterium colony count results are 28 ± 7.
(3) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, the composition of the levels of double-layer plate all containing N-acetylcystein and chloramphenicol, it is combined as 1~5mg/L N-
Acetylcysteine+10mg/L chloramphenicol), good double-layer plate is placed in 37 DEG C of constant incubator quiescent culture 36h simultaneously
Count.Bacterium colony count results are 0.
Embodiment 4：A kind of combined method for improving chloramphenicol effect of drugs, comprises the following steps：
(1) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate is placed in quiescent culture 36h in 37 DEG C of constant incubator (double-layer plate is free of chloramphenicol), counts E. coli clones number,
As a control group.Bacterium colony count results are 59 ± 5.
(2) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, in double-layer plate containing chloramphenicol and levels concentration it is consistent and be 10mg/L, good double-layer plate is placed in 37 DEG C of constant temperature
Quiescent culture 36h and counted in incubator.Bacterium colony count results are 28 ± 7.
(3) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, the composition of the levels of double-layer plate all containing N-acetylcystein and chloramphenicol, it is combined as 6~10mg/L
N-acetylcystein+10mg/L chloramphenicol), good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator
And count.Bacterium colony count results are 0.
Embodiment 5：A kind of combined method for improving chloramphenicol effect of drugs, comprises the following steps：
(1) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate is placed in quiescent culture 36h in 37 DEG C of constant incubator (double-layer plate is free of chloramphenicol), counts E. coli clones number,
As a control group.Bacterium colony count results are 59 ± 5.
(2) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, in double-layer plate containing chloramphenicol and levels concentration it is consistent and be 10mg/L, good double-layer plate is placed in 37 DEG C of constant temperature
Quiescent culture 36h and counted in incubator.Bacterium colony count results are 31 ± 8.
(3) the Escherichia coli dilution 10 of logarithmic phase5Times, take the Escherichia coli after 50 μ L dilutions to fall as stated above double-deck flat
Plate, the composition of the levels of double-layer plate all containing N-acetylcystein and chloramphenicol, it is combined as 11~15mg/L
N-acetylcystein+10mg/L chloramphenicol), good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator
And count.Bacterium colony count results are 0.
Although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
A variety of changes, modification can be carried out to these embodiments, replace without departing from the principles and spirit of the present invention by understanding
And modification, the scope of the present invention is defined by the appended.
Claims (1)
1. a kind of combined method for improving chloramphenicol effect of drugs, it is characterised in that comprise the following steps：
A. chloramphenicol is prepared：The chloramphenicol mother liquor for being 20mg/mL with absolute ethyl alcohol compound concentration, and with 0.22 μm of membrane filtration
It is degerming, it is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of b.N- acetylcysteines：10mg/mL N-acetylcystein is prepared with deionized water and with 0.22 μm
Membrane filtration is degerming, is placed in after packing standby in -20 DEG C of refrigerator；
The preparation of c.LB fluid nutrient mediums：Sodium chloride 5g, peptone 10g, dusty yeast 5g, 1000mL is added water to, pH is naturally, packing
After 1.034 × 105Pa sterilizing 20min, solid medium add w/v 1.2% agar powder thereto；Semisolid culturemedium
Add w/v 0.5% agar powder；
D. the solid substrate flat board of chloramphenicol is free of：LB solid mediums after sterilizing pour into nothing after it is cooled to 50 DEG C or so
The disposable plate of bacterium, cooling are stand-by；
Bottom solid plate containing chloramphenicol：LB solid mediums after sterilizing add thereto after it is cooled to 50 DEG C or so
Chloramphenicol mother liquor pours into sterile disposable plate, cooling is stand-by to the working concentration needed after mixing；
E. the culture of Escherichia coli：E. coli broth is inoculated with to fresh LB fluid nutrient mediums according to the inoculum concentrations of w/v 2%
In cultivated in 37 DEG C of 180rpm to logarithmic phase, it is stand-by；
F. the dilution of Escherichia coli：The Escherichia coli of step e culture are taken to dilute 105It is stand-by again；
G. semi-solid upper panel is fallen without chloramphenicol：50 μ L are taken to dilute 105Escherichia coli again are added in ampere bottle, then to
The semisolid of 5mL sterilizings is added in ampere bottle, semi-solid temperature is poured on solid plate good in advance at 45 DEG C after shaking up；
Semi-solid upper panel is fallen containing chloramphenicol：50 μ L are taken to dilute 105Times Escherichia coli be added in ampere bottle, then to ampere
The semisolid of 5mL sterilizings is added in bottle, semi-solid temperature adds chloramphenicol at 45 DEG C, and in semisolid to the concentration needed, shakes
Poured into after even on the solid plate of the advance good chloramphenicol containing same concentrations；
H. good double-layer plate is placed in quiescent culture 36h in 37 DEG C of constant incubator, bacterium colony is counted.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201710928120.1A CN107648214A (en) | 2017-09-22 | 2017-09-22 | A kind of combined method for improving chloramphenicol effect of drugs |
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| CN201710928120.1A CN107648214A (en) | 2017-09-22 | 2017-09-22 | A kind of combined method for improving chloramphenicol effect of drugs |
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| CN201710928120.1A Pending CN107648214A (en) | 2017-09-22 | 2017-09-22 | A kind of combined method for improving chloramphenicol effect of drugs |
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2017
- 2017-09-22 CN CN201710928120.1A patent/CN107648214A/en active Pending
Non-Patent Citations (2)
| Title |
|---|
| MANISH GOSWAMI等: "Antibiotics and Antioxidants: Friends or Foes During Therapy?", 《BARC NEWSLETTER》 * |
| 张亚妮等: "N-乙酰半胱氨酸对铜绿假单胞菌的耐药性及绿脓菌素产量的影响", 《中国药理学通报》 * |
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Application publication date: 20180202 |