阅读说明：本技术 色氨酸增强对革兰氏阴性菌杀菌作用的应用 (Use of tryptophan for enhancing gram-negative bacteria bactericidal effect ) 是由 李艳 张利宁 刘巍峰 丛华 郭晶晶 孙夏慧 于 2018-07-12 设计创作，主要内容包括：本发明公开了色氨酸具有增强革兰氏阴性菌杀菌效果的作用,具体涉及色氨酸在制备革兰氏阴性菌杀菌药物中的应用。本发明设计实验证明了庆大霉素和卡那霉素中添加色氨酸能够提升细菌胞内活性氧水平,同时还能够提升细菌对于抗生素的吸收从而改善细菌对药物的敏感性,充分验证了色氨酸对革兰氏阴性菌的诱杀作用以及对耐药菌的杀菌效果的提升。基于以上发现,本发明相应的保护色氨酸的第二医疗用途、一种杀菌组合物以及一种杀菌药物以及一种杀菌方法。(The invention discloses an effect of tryptophan on enhancing gram-negative bacteria sterilization effect, and particularly relates to an application of tryptophan in preparation of gram-negative bacteria sterilization medicines. The design and experiments of the invention prove that the level of intracellular active oxygen of bacteria can be improved by adding tryptophan into gentamicin and kanamycin, and the absorption of bacteria to antibiotics can be improved, so that the sensitivity of bacteria to drugs can be improved, and the trapping and killing effects of tryptophan on gram-negative bacteria and the improvement of the sterilization effect of drug-resistant bacteria can be fully verified. Based on the above findings, the invention provides a second medical application of the corresponding protected tryptophan, a bactericidal composition, a bactericidal medicament and a bactericidal method.)

1. Application of tryptophan in preparing medicine for killing gram-negative bacteria is provided.

2. Use according to claim 1, characterized in that said gram-negative bacteria are susceptible escherichia coli and resistant escherichia coli.

3. Use according to claim 2, wherein the drug-resistant E.coli is an aminoglycoside-resistant E.coli.

4. The use of claim 3, wherein the aminoglycoside is gentamicin and kanamycin.

5. A bactericidal composition against gram-negative bacteria, characterized by comprising an antibiotic and tryptophan as ingredients.

6. The bactericidal composition of claim 5, wherein the antibiotic is an aminoglycoside antibiotic.

7. The bactericidal composition of claim 6, wherein the aminoglycoside antibiotic is gentamicin or kanamycin.

8. A bactericidal medicine for gram-negative bacteria, which is characterized in that the specific components of the bactericidal medicine are the bactericidal composition in claims 5-7, and pharmaceutically acceptable auxiliary materials and carriers.

9. A method for killing gram-negative bacteria, wherein the medicament used in the method is a preparation prepared from the bactericidal composition of claim 8 and pharmaceutically acceptable auxiliary materials.

Technical Field

The invention relates to the technical field of gram-negative bacteria killing, in particular to application of tryptophan capable of enhancing antibiotic killing of gram-negative bacteria.

Background

With the widespread use of antibiotics, more and more resistant and resistant bacteria are emerging. This makes it extremely difficult to treat bacterial infections in the future. Drug-resistant bacteria are in turn a significant cause of chronic and recurrent infections, such as urinary tract infections. The results of the current studies indicate that 90% of chronic urinary tract infections are caused by e. However, the research and development period of the novel antibiotics is longer, and bacteria have more adaptive mechanisms along with the change of environment, so that the safe and effective bactericidal enhancer is a very promising effective way for treating bacterial infection. If the effect of enhancing both resistant bacteria and drug-resistant bacteria can be well enhanced, the method has great clinical application prospect.

Tryptophan is one of 8 amino acids essential to human body, and has functions of relieving insomnia and stabilizing emotion, and has effects in brain together with vitamin B6, nicotinic acid and magnesium, and can be used for producing serotonin in blood, which is an essential nerve activity conducting substance and neurohormone for making human sleep normally, and plays an important role in maintaining various physiological functions of human body. In addition, tryptophan can be used as additive for improving mental state and reducing depression. Therefore, tryptophan is a safer and more reliable way to be used as a bactericidal enhancer.

Disclosure of Invention

The invention verifies that the tryptophan can promote sterilization by improving the level of Reactive Oxygen Species (ROS) in cells, and the tryptophan can improve the sensitivity of bacteria to antibiotics to kill resistant bacteria and reduce the generation effect of drug-resistant bacteria.

Based on the experimental conclusion, the invention provides the application of tryptophan in the preparation of the bactericidal medicine, and provides a bactericidal composition, a bactericidal medicine and a bactericidal method.

The invention aims to provide application of tryptophan in preparation of a bactericidal medicament, which is characterized in that the bactericidal medicament is a medicament for gram-negative bacteria.

Preferably, the gram-negative bacteria are susceptible E.coli and drug-resistant E.coli.

Further, the drug-resistant Escherichia coli is an aminoglycoside-resistant Escherichia coli.

Further, the bactericidal drugs are gentamicin and kanamycin.

The invention also provides a bactericidal composition which is characterized by comprising the components of the combination of antibiotics and tryptophan.

Preferably, the antibiotic is an aminoglycoside antibiotic.

Further, the aminoglycoside antibiotic is gentamicin or kanamycin.

The invention also provides a bactericidal medicine, which aims at gram-negative bacteria and is characterized in that the specific component of the bactericidal medicine is one of the bactericidal compositions.

The invention also provides a method for killing gram-negative bacteria, which is characterized in that the medicament used in the method is a preparation prepared from the bactericidal composition and pharmaceutically acceptable auxiliary materials.

The invention has the advantages of

1. After the bacteria obtain the drug resistance, the treatment effect of the chemical drug is greatly reduced, and meanwhile, the drug resistance gene can be inherited through the plasmid to become the inherent drug-resistant bacteria. The tryptophan and the antibiotics can obtain better treatment effect, reduce the dosage of chemical drugs, reduce the generation of drug-resistant bacteria, and provide a new idea for the research and development of bactericidal drugs.

2. Tryptophan is used as amino acid necessary for human body, and has high organism acceptance, small toxic and side effects and low research and development cost.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this application, illustrate embodiments of the application and, together with the description, serve to explain the application and are not intended to limit the application.

FIG. 1 is a graph of the survival rate of susceptible and resistant bacteria treated with antibiotic in combination with tryptophan;

(a) the method comprises the following steps Bacterial survival results for MG1655 at logarithmic growth phase at various concentrations of gentamicin (0MG/L,1MG/L,2MG/L,3MG/L,4MG/L) and with and without 5mM tryptophan for twenty-four hours.

(b) MG1655 at logarithmic growth phase survived twenty-four hours after treatment with different concentrations of kanamycin (0MG/L,0.5MG/L,1.5MG/L,2.5MG/L,3.5MG/L) and with or without 5mM tryptophan.

(c) In the kanamycin treatment of 2mg/L or 1.5mg/L gentamicin with the concentration lower than the minimum inhibitory concentration, tryptophan with different concentrations is added in sequence, and the result of the bacterial survival rate is measured.

(d) Stationary MG1655 bacterial survival after one hour of gentamicin 60MG/L or kanamycin 40MG/L and with or without 5mM tryptophan treatment.

(e) The bactericidal effect of tryptophan on resistant bacteria. The viability of the bacteria was 24 hours after 24 hours of logarithmic growth phase EY1 treated with gentamicin 60mg/L or kanamycin 40mg/L alone or in combination with 5mM tryptophan.

FIG. 2 is a graph showing the effect of tryptophan on the production rate of drug-resistant bacteria;

(a) the ratio of drug-resistant bacteria production was determined by inoculating 1% of the overnight culture to a fresh medium, culturing MG1655 at different times (from 3 hours to 9 hours) for 5 hours after treatment with 60MG/L gentamicin, and adding tryptophan at different concentrations for 30 minutes.

(b) The ratio of drug-resistant bacteria production was determined by inoculating overnight cultures to fresh medium at 1% transfer rate, treating MG1655 cultured for different periods of time (from 3 hours to 9 hours) with kanamycin at 40MG/L for 5 hours, and then treating for 30 minutes with tryptophan at different concentrations.

(c) The bactericidal effect of tryptophan on drug-resistant bacteria. MG1655 in logarithmic phase was treated with gentamicin 60MG/L or kanamycin 40MG/L for 5 hours, and the cells were centrifuged, washed once with 1 × PBS buffer, and centrifuged. The viable bacteria count is divided into two parts, one part is directly counted, the other part is added with antibiotics for independent treatment or the antibiotics and 5mM tryptophan are jointly treated for five hours, and the proportion of the bacteria before and after treatment is compared.

FIG. 3 is a graph showing the results of measurement of intracellular reactive oxygen species of bacteria

MG1655 in logarithmic and stationary phase was treated with gentamicin at 60MG/L alone or in combination with 5mM tryptophan for one hour to determine intracellular reactive oxygen species levels.

FIG. 4 is a graph of the results of antibiotic uptake assays.

(a) Uptake of Texas Red-labeled gentamicin by MG1655 at logarithmic growth phase at different concentrations of tryptophan.

(b) Uptake of Texas Red-labeled gentamicin by stationary-phase MG1655 at different concentrations of tryptophan.

Detailed Description

It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.