Antibiotic Adjuvant and Its Preparation Method and Application

Abstract

The present invention provides an antibiotic adjuvant, its preparation method and application, specifically provides an antimicrobial peptide FP-CATH9 derived from Deinagkistrodon acutus and its application in antibacterial sensitization. FP-CATH9 itself does not have antibacterial activity, but it has the characteristics of small molecular weight, simple synthesis, low hemolytic activity, serum tolerance, good heat tolerance and thermal stability, and can be applied to the fields of medicine, cosmetics, food preservation and aquaculture. The present invention expands the application of cationic short peptides without antibacterial activity and provides new insights and examples for the research and development of antibiotic sensitizers.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and particularly to an antibiotic adjuvant, a preparation method thereof, and an application thereof. Background Art

[0002] Tetracycline antibiotics include chlortetracycline, tetracycline, minocycline, tigecycline, etc. They form reversible binding complexes with the 30S subunit of ribosomes in bacteria to inhibit protein synthesis and achieve antibacterial effects. Although tigecycline has strong antibacterial effects, it is expensive. Compared with other tetracycline antibacterial drugs, minocycline has higher lipid solubility and some significant characteristics in pharmacokinetics, specifically in the following aspects: strong tissue permeability, high bioavailability, long half-life, antibacterial activity of metabolites, and better tolerance in patients with renal insufficiency. Traditional antibiotics have achieved remarkable effects in the treatment of bacterial infectious diseases. However, in recent years, with the abuse of traditional antibiotics in the fields of medicine and aquaculture, etc., microorganisms have developed stronger and stronger tolerance to traditional antibiotics, and the problem of microbial drug resistance has become a difficult problem seriously threatening human health. At present, the drug resistance rates of Klebsiella pneumoniae and Acinetobacter baumannii to minocycline and tigecycline are increasing, while Pseudomonas aeruginosa is naturally resistant to tetracycline drugs. The research and development of antibiotics is difficult, and no original antibiotics have been marketed for nearly 30 years. Antibiotic sensitizers are promising antibacterial adjuvants. Such drugs are rarely used alone because they have poor antibacterial effects or no antibacterial effects when used alone. Trimethoprim is a good example. When used in combination with antibiotics, it has obvious synergistic effects and can increase by several times to hundreds of times.

[0003] Antibacterial peptides are small-molecule polypeptides that have killing effects on bacteria, fungi, viruses, protozoa, etc. Antibacterial peptides have the characteristics of small molecular weight, simple structure, strong bactericidal activity, etc. Moreover, antibacterial peptides generally have no toxicity to normal mammalian cells and tissues, have no residue problems, and have good biocompatibility. However, some truncated parts of natural antibacterial peptides have no antibacterial activity, and the functions of these short peptides have not been fully and detailedly elucidated. Summary of the Invention

[0004] Therefore, the present invention provides an antibiotic adjuvant, a preparation method thereof, and an application thereof. The inventor combines the truncated peptide FP-CATH9 (which itself has no antibacterial effect) derived from the Cathelicidin antibacterial peptide FP-CATH of Deinagkistrodon acutus with minocycline, and shows extremely strong effects against minocycline-resistant Gram-negative bacteria (such as Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, etc.). This invention expands the application of cationic short peptides that have no antibacterial effect and provides new insights and examples for the research and development of antibiotic sensitizers.

[0005] An antibiotic adjuvant, which is derived from the truncated peptide FP-CATH9 of the Cathelicidin antimicrobial peptide FP-CATH of Deinagkistrodon acutus, and the antibiotic adjuvant consists of 9 amino acids, and the amino acid sequence is:

[0006] Lys Arg Phe Lys Lys Phe Trp Lys Lys, and all amino acids are L-type.

[0007] A preparation method of an antibiotic adjuvant, based on obtaining the amino acid sequence FP-CATH of the mature peptide of Deinagkistrodon acutus to obtain the FP-CATH9 sequence, synthesizing its full sequence with an automatic polypeptide synthesizer, and desalting and purifying it by HPLC reverse-phase column chromatography; identifying its purity by high-performance liquid chromatography (HPLC) method, identifying the sequence by LC-MS / MS, determining the isoelectric point by isoelectric focusing electrophoresis, determining the secondary structure by circular dichroism spectroscopy, and predicting the tertiary structure of the amino acid sequence by SWISS software.

[0008] The application of the above-mentioned antibiotic adjuvant, which is used for preparing any one of antibacterial drugs or compositions, drugs or compositions for inhibiting bacterial growth, preservatives, animal feed additives, and cosmetic additives.

[0009] According to the application of the above-mentioned antibiotic adjuvant, the antibiotic adjuvant is used in combination with tetracycline antibiotics.

[0010] According to the application of the above-mentioned antibiotic adjuvant, the tetracycline antibiotic is minocycline or tigecycline.

[0011] The antibiotic adjuvant provided by the present invention, its preparation method and application provide an antimicrobial peptide FP-CATH9 derived from Deinagkistrodon acutus and its application in antibacterial sensitization. FP-CATH9 itself does not have antibacterial activity, but it has the characteristics of small molecular weight, simple synthesis, low hemolytic activity, serum tolerance, heat tolerance and good thermal stability, and can be applied to the fields of medicine, cosmetics, food preservation and aquaculture. The present invention expands the application of cationic short peptides without antibacterial activity and provides new insights and examples for the research and development of antibiotic sensitizers. Description of the Drawings

[0012] The above-mentioned and / or additional aspects and advantages of the embodiments of the present invention will become obvious and easy to understand from the description of the embodiments in conjunction with the following drawings, where:

[0013] Figure 1 It is a schematic diagram of the sensitization effect of FP-CATH9 on minocycline and tigecycline, and the test bacteria: Klebsiella pneumoniae 3 resistant to minocycline;

[0014] Figure 2It is a schematic diagram of the checkerboard method for detecting the synergistic effect of FP-CATH9 and minocycline. Test bacteria: Klebsiella pneumoniae 3 resistant to minocycline.

[0015] Figure 3 It is a schematic diagram for detecting the sensitization effect of FP-CATH9 on minocycline and tigecycline, with Pseudomonas aeruginosa ATCC 27853 (A) and Acinetobacter baumannii (B) resistant to minocycline clinically as test bacteria;

[0016] Figure 4 It is a schematic diagram of the bactericidal speed of FP-CATH9 sensitizing minocycline. Test bacteria: Klebsiella pneumoniae 3 resistant to minocycline.

[0017] Figure 5 It is a schematic diagram for the determination of the hemolytic activity of human red blood cells by different concentrations of FP-CATH9 and in combination with minocycline;

[0018] Figure 6 It is a schematic diagram for the determination of the cytotoxicity of different concentrations of FP-CATH9 and in combination with minocycline on mouse Raw264.7 cells;

[0019] Figure 7 It is a schematic diagram of the protective effect of different concentrations of FP-CATH9 and in combination with minocycline on Galleria mellonella. Detailed implementation manners

[0020] For ease of understanding the present invention, the present invention will be described more comprehensively below with reference to each embodiment. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, these embodiments are provided to make the disclosure of the present invention more thorough and comprehensive.

[0021] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the technical field to which the present invention belongs. The terms used in the specification of the present invention herein are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" used herein includes any and all combinations of one or more of the related listed items.

[0022] The embodiments of the present invention will be further described below in multiple embodiments. The embodiments of the present invention are not limited to the following specific embodiments. Within the scope of the main rights unchanged, appropriate changes can be made for implementation.

[0023] Example 1:

[0024] An antibacterial peptide FP-CATH9 from Deinagkistrodon acutus

[0025] A gene sequence (FP-CATH) was found by retrieving the genome data of Deinagkistrodon acutus from the NCBI website. The mature peptide has a length of 34 amino acids. FP-CATH9 is a truncated peptide sequence of FP-CATH, with a molecular weight of 1295.64 and an isoelectric point of 11.39. The sequence listing SEQ ID NO:1 is as follows:

[0026] Lys Arg Phe Lys Lys Phe Trp Lys Lys, and all amino acids are of the L-type.

[0027] Example 2

[0028] A chemical synthesis method for the short peptide FP-CATH9 derived from the antimicrobial peptide of Deinagkistrodon acutus, comprising:

[0029] 1. First, synthesize FP-CATH9, and desalt and purify it by HPLC-C 18 reverse-phase column chromatography.

[0030] 2. The molecular weight is determined by LC-MS / MS method.

[0031] 3. The purified FP-CATH9 is identified by high-performance liquid chromatography (HPLC) with a purity > 95%, a molecular weight of 1295.64, and an isoelectric point of 11.39 determined by isoelectric focusing electrophoresis.

[0032] Example 3:

[0033] Antibacterial activity detection of the short peptide FP-CATH9

[0034] (1) Respectively pick the activated test strain single colonies (such as Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, clinically isolated minocycline-resistant Klebsiella pneumoniae, and Acinetobacter baumannii ATCC 19606, etc., which are preserved in the laboratory), dilute them with normal saline to a concentration of 0.5 McFarland units, and evenly coat them on the MH solid medium (purchased from Shanghai Guandao Bioengineering Co., Ltd.) plate with a sterile cotton swab. Place a sterilized filter paper with a diameter of 6 mm on the surface of the medium, and drop 10 μL of the 3.2 mg / mL FP-CATH9 sample solution dissolved in normal saline. Incubate it inverted at 37 °C for 18 - 20 h, and observe whether an inhibition zone is formed. If the sample has antibacterial activity, a clear and transparent inhibition zone will be formed around the filter paper, and the larger the inhibition zone, the stronger the antibacterial activity of the sample.

[0035] (2) The minimum inhibitory concentration (MIC) was determined by the double dilution method: The strains in the above experiment (1) were selected for the MIC determination experiment. The test strains were inoculated into MH liquid medium (purchased from Shanghai Guandao Biotechnology Co., Ltd.), and then cultured with shaking at 37 °C in an incubator until the logarithmic growth phase. Then, the above strain culture solution was diluted to 1.5×10 8 cfu / mL for use. First, 10 μL of MH liquid medium was added to each well of a sterile 96-well plate, and then 10 μL of the FP-CATH9 sample solution (filtered through a 0.22 μm pore filter membrane) was aspirated and sequentially diluted one by one in order. 10 μL was aspirated and discarded from the 11th well, and the 12th well was the negative control well. 90 μL of the prepared bacterial solution (the bacterial solution concentration was 200 times the dilution of the above mother bacterial solution) was added to each well, and then the 96-well plate was placed in an incubator and statically cultured at 37 °C for 18 h. The optical absorption was measured at a wavelength of 600 nm. The minimum inhibitory concentration was the lowest sample concentration at which no bacterial growth was visible.

[0036] As can be seen from Table 1, FP-CATH9 did not show antibacterial activity against the above bacteria (except that the MIC value for Escherichia coli ATCC25922 was 128 μg / mL).

[0037] Table 1

[0038]

[0039] Example 4:

[0040] Screening of the antibacterial and sensitizing activity of the short peptide FP-CATH9 against antibiotics

[0041] Respectively pick single colonies of the well-activated test strains (Klebsiella pneumoniae 3 resistant to minocycline), dilute them with normal saline to a concentration of 0.5 McFarland units, and evenly coat them on the MH solid medium (purchased from Shanghai Guandao Biotechnology Co., Ltd.) plates. Take two drug sensitivity test papers containing the same antibiotic each (from Thermo Fisher Scientific) and place them on the surface of the medium. Then, respectively add 10 μL of FP-CATH9 solution (dissolved in normal saline, with a concentration of 3.2 mg / mL) and 10 μL of normal saline. On the same plate, simultaneously stick antibiotic papers such as IPM, MEM, CIP, SXT, MH, TGC, AK, CRO, CAZ, GN, AMP, and TZP, and incubate them upside down at 37 °C for 18 - 20 h, and observe whether there are differences in the formation of inhibition zones and the normal saline of their respective drugs. If the sample has antibacterial activity, a clear and transparent inhibition zone will be formed around the filter paper. An increase in the inhibition zone compared to the normal saline group indicates a possible antibacterial sensitization effect. The results show that FP-CATH9 does not have sensitization activity against the antibiotics listed in Table 2 (except minocycline and tigecycline). 10 μL of FP-CATH9 can sensitize the minocycline inhibition zone from 6 mm to 15 mm, and sensitize the tigecycline inhibition zone from 16 mm to 18 mm. The results are shown in Figure 1 。

[0042] Table 2

[0043]

[0044] Example 5:

[0045] Antibacterial sensitization activity of the short peptide FP-CATH9 against minocycline

[0046] (1) Checkerboard experiment

[0047] Take the above-mentioned Klebsiella pneumoniae 3 strains in the logarithmic phase (cultured with shaking at 37 °C), and dilute them with fresh MH liquid medium to 1.5×10 8 cfu / mL for standby. Use a 96-well sterile microplate. The highest concentration of minocycline is 128 μg / mL, and the highest concentration of FP-CATH is 256 μg / mL. Each antibacterial drug is serially diluted 2-fold with sterilized MH broth from the highest concentration in sequence. Take 10 μL of each and arrange them in the rows and columns of the plate respectively. Then, add 80 μL of the bacterial solution (the bacterial solution concentration is the mother solution diluted 200-fold as above) to the sterile microplate, and fill the other wells with normal saline to a volume of 100 μL. Incubate them upright at 37 °C for 18 - 20 h. The lowest drug concentration without bacterial growth is the MIC. The checkerboard experiment for combined antibacterial activity refers to the operating procedures of the Clinical and Laboratory Standards Institute (CLSI) of the United States. The fractional inhibitory concentration index (FICI) is used for the combined drug sensitivity experiment by the microbroth checkerboard dilution method. The FICI index value is calculated as follows: FICI = MIC ab / MICa +MIC ba / MIC b 。

[0048] The FICI index ≤ 0.5 indicates synergy; 0.5 < FIC ≤ 1 indicates additivity; 1 < FIC ≤ 2 indicates no interaction; FIC > 2 indicates antagonism.

[0049] Figure 2 The results showed that the FICI index of FP-CATH9 and minocycline was 0.0195, indicating a strong synergistic effect.

[0050] (2) Plate experiment

[0051] Separate single colonies of the activated test strains (Pseudomonas aeruginosa ATCC 27853 and Acinetobacter baumannii from clinical samples) were picked and diluted to a concentration of 0.5 McFarland units with normal saline. They were evenly spread on the surface of MH solid medium (purchased from Shanghai Guandao Bioengineering Co., Ltd.) plates using a sterile cotton swab. Two minocycline susceptibility test discs (Thermo Fisher Scientific) were placed on the surface of the medium. 10 μL of a 3.2 mg / mL FP-CATH9 sample solution dissolved in normal saline was added dropwise to one disc, and 10 μL of normal saline was added dropwise to the other disc. The plates were incubated inverted at 37°C for 18 - 20 h, and the formation of inhibition zones was observed. If the sample had antibacterial activity, a clear and transparent inhibition zone would form around the filter paper disc. An increase in the inhibition zone compared to the normal saline group indicated possible antibacterial sensitization. 10 μL of EDTA (0.2 mol / L) was used as a positive control. Figure 3 A shows that for Pseudomonas aeruginosa ATCC 27853, FP-CATH9 showed extremely strong sensitizing activity against both minocycline and tigecycline. Although EDTA also showed a sensitizing effect, upon careful observation, the transparency of the inhibition zone was not as high as that of the FP-CATH9 group, indicating weak bacterial growth. Figure 3 B shows that for Acinetobacter baumannii 2 from clinical samples (resistant to minocycline and intermediate to tigecycline), FP-CATH9 showed extremely strong sensitizing activity against minocycline and also had a certain sensitizing effect on tigecycline. In summary, the results indicate that FP-CATH9 has sensitizing activity against both minocycline and tigecycline.

[0052] Example 6:

[0053] Determination of the bactericidal rate enhancement of the short peptide FP-CATH9 on minocycline

[0054] Take the above-mentioned Klebsiella pneumoniae 3 strain in the logarithmic phase (cultured with shaking at 37°C) and dilute it with fresh MH liquid medium to 10 6Bacterial suspension of cfu / mL. FP-CATH9, minocycline, and FP-CATH9 + minocycline dissolved in physiological saline were added to the bacterial suspension to a final concentration of (16 μg / mL) for both. The bacterial suspension with the added drugs was placed in an incubator and shaken at 37 °C. At 0, 2, 4, 6, 8, 10, and 12 h, 50 μL of the bacterial suspension was diluted 1000-fold, and then 50 μL of the diluted bacterial suspension was spread on MH solid medium and cultured overnight at 37 °C for colony counting. Physiological saline was used as a negative control.

[0055] The results are as Figure 4 shown. The combination of FP-CATH9 (16 μg / mL) + minocycline (16 μg / mL) could completely kill the bacteria at 10 h. However, the treatment of bacteria with FP-CATH9 (16 μg / mL) and minocycline (16 μg / mL) returned to the growth level of the physiological saline group after 10 h, indicating no antibacterial function.

[0056] Example 7:

[0057] (1) Determination of hemolytic activity

[0058] The standard human blood cell reagent (Changchun Boxun Biotechnology Co., Ltd.) was washed 3 times with physiological saline and prepared into a red blood cell suspension of 10 7 cell / mL. The above red blood cell suspension was mixed with a certain concentration of FP-CATH9, FP-CATH9 + minocycline, and minocycline dissolved in physiological saline, incubated at 37 °C for 1 h, then centrifuged at 1000 rpm for 5 min, the supernatant was aspirated, and the absorbance was measured at a wavelength of 540 nm. Physiological saline was used as a negative control, and Triton X-100 was used as a positive control.

[0059] The percentage of hemolysis rate was calculated according to the following formula: Hemolysis rate % = A 样品 - A 阴性对照 / A 阳性对照 × 100%.

[0060] The results Figure 5 showed that FP-CATH9 had low hemolytic activity. The hemolytic activity of the combination of FP-CATH9 (128 μg / mL) and minocycline (128 μg / mL) was about 15%, and its concentration far exceeded its effective bactericidal concentration, indicating that FP-CATH9 had great application potential.

[0061] (2) Determination of cytotoxicity

[0062] The cytotoxicity of the drug on mouse Raw264.7 cells was determined by the CCK method. FP-CATH and minocycline were combined and arranged at different final concentrations. The specific steps are briefly as follows: The drug was dissolved in serum-free RPMI 1640 medium and then added to a 96-well plate containing mouse Raw264.7 cells (2×10 4 cells / well), and serum-free RPMI 1640 medium without the drug was used as a blank control. After incubation for 24 h, 10 μL of CCK 8 reagent was added to each well. After incubation for 4 h, the absorbance at 450 nm was measured. For 3 independent experiments, the results were averaged, and the cell viability % was calculated as = [A 加药 -A 空白 / [A 0加药 -A 空白 ×100%. A 空白 represents the absorbance of the well containing medium and CCK solution but no cells. The results Figure 6 showed that FP-CATH9 had low hemolytic activity. The hemolytic activity of the combination of FP-CATH9 (128 μg / mL) and minocycline (128 μg / mL) was about 15%, and its concentration far exceeded its effective bactericidal concentration, indicating that FP-CATH9 had great application potential.

[0063] Example 8:

[0064] Serum stability, heat tolerance and thermal stability of the short peptide FP-CATH9

[0065] (1) Pick a single colony of the activated test strain (Klebsiella pneumoniae 3 strain clinically), dilute it with normal saline to a concentration of 0.5 McFarland units, and evenly coat it on the surface of an MH solid medium (purchased from Shanghai Guandao Biotechnology Co., Ltd.) plate with a sterile cotton swab. Place two minocycline susceptibility test discs (Thermo Fisher Scientific) on the surface of the medium. According to serum: FP-CATH9 (C:C), at concentrations of 0:1, 0.125:1, 0.25:1, 0.5:1, 1:1, 1:0 respectively, a total of 10 μL was added dropwise to the minocycline susceptibility test discs. As shown in Table 4. The plate method was used to determine the effect of serum on the sensitization of FP-CATH9. Add 5 μL of the 6.4 mg / mL FP-CATH9 sample solution dissolved in normal saline to one disc, and add 5 μL of serum dilution or normal saline to the other disc. Incubate at 37 °C in an inverted position for 18 - 20 h, and observe whether an inhibition zone is formed. If the sample has antibacterial activity, a clear and transparent inhibition zone will form around the filter paper disc. Compare the changes in the size of the inhibition zone with the normal saline group and the serum dilution group. The results are shown in Table 3. When FP-CATH9 (C:C) was 1:1, the sensitization effect decreased slightly, but the inhibition zone was still 11 mm compared with 6 mm in the blank group.

[0066] Table 3

[0067]

[0068] (2) Determination of the thermal tolerance of FP-CATH9: Dissolve FP-CATH9 in sterilized deionized water to prepare a sample solution with a concentration of 3.2 mg / mL. Incubate it in a constant-temperature metal bath at 4 °C, 20 °C, 40 °C, 60 °C, and 80 °C for 1 h each. After cooling, take 10 μL of the incubated FP-CATH9 sample solution and use the plate method to determine the sensitization effect of FP-CATH9 on Klebsiella pneumoniae 3 resistant to minocycline. The results are shown in Table 4. FP-CATH9 still maintains a high sensitization activity after treatment at 60 °C and 80 °C.

[0069] Table 4

[0070]

[0071] (3) Determination of the thermal stability of FP-CATH9: Dissolve FP-CATH9 in sterilized deionized water to prepare a sample solution with a concentration of 3.2 mg / mL. Incubate it in an incubator at 37 °C for 0 - 48 h. Take the FP-CATH9 sample solution at 0, 12, 24, and 48 h respectively, and use the plate method to determine the sensitization effect of FP-CATH9 on Klebsiella pneumoniae 3 resistant to minocycline. The results are shown in Table 5. FP-CATH9 still maintains a high sensitization activity after treatment at 24 and 48 h.

[0072] Table 5

[0073]

[0074] Example 9: In vivo activity determination experiment

[0075] (1) Construction of a Galleria mellonella infection model

[0076] Preparation of the strain infection sample: Pick a single colony of Klebsiella pneumoniae 3 resistant to minocycline and place it in 5 mL of LB broth. Place it in a constant-temperature shaker at 200 rpm / min and 37 °C for 12 h. After 12 h, expand the culture, dilute it at a ratio of 1:100, take the logarithmic-phase bacterial solution, wash it with physiological saline and resuspend it in a 1 mL system to prepare a bacterial solution concentration of 1.5×10 8 cfu / mL. The lethal concentration has been explored in advance. Randomly divide the Galleria mellonella into five groups: inject Klebsiella pneumoniae 3 resistant to minocycline (treated group), single administration of FP-CATH (16 μg / mL) group, single administration of minocycline (16 μg / mL) group, FP-CATH (16 μg / mL) + minocycline (16 μg / mL) group, and NS group (physiological saline group).

[0077] (2) Survival experiment of Galleria mellonella

[0078] Absorb 10 μL of the bacterial solution of Klebsiella pneumoniae strain 3 resistant to minocycline (concentration: 1.5×10 8 cfu / mL). Immediately after mixing the drug and the bacterial solution evenly, inject it into Galleria mellonella. Use a micro syringe to inject it into the right hind limb of the larva, and incubate the infected larva in the dark at 37 °C without food for up to 7 days. For each group of Galleria mellonella (n = 10), monitor the survival rate after the Galleria mellonella is infected, record the survival of Galleria mellonella every 24 h, and define it as dead when they have no response to touch. The results are as Figure 7 shown. The survival rate of the normal saline group (NS) in 7 days is 100%. All the Galleria mellonella injected with Klebsiella pneumoniae strain 3 resistant to minocycline (treated group) died within 1 day. FP-CATH (16 μg / mL) and minocycline (16 μg / mL) alone basically have no protective effect. While the survival rate of the FP-CATH (16 μg / mL) + minocycline (16 μg / mL) group in 7 days is 80%, showing a strong protective effect.

[0079] It can be seen from the above embodiments that FP-CATH9 in the present invention can be obtained by chemical synthesis. This peptide has extremely strong sensitizing activity against Klebsiella pneumoniae resistant to minocycline, Acinetobacter baumannii, and Pseudomonas aeruginosa (naturally resistant to minocycline and tigecycline), including some clinically isolated pathogenic bacteria. Secondly, this antimicrobial peptide has the characteristics of small molecular weight, simple synthesis, low hemolytic activity, serum tolerance, good heat tolerance and thermal stability, and can be applied to the fields of medicine, cosmetics, food preservation and aquaculture.

[0080] Although the embodiments of the present invention have been shown and described, those of ordinary skill in the art can understand that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principles and purposes of the present invention. The scope of the present invention is defined by the claims and their equivalents.

Claims

1. An antibiotic adjuvant, characterized in that the antibiotic adjuvant is derived from the truncated peptide FP-CATH9 of the Cathelicidin antimicrobial peptide FP-CATH of Deinagkistrodon acutus, and the antibiotic adjuvant consists of 9 amino acids, and the amino acid sequence is: Lys Arg Phe Lys Lys Phe Trp Lys Lys, and all amino acids are of the L-type.

2. A method for preparing the antibiotic adjuvant according to claim 1, characterized in that based on the amino acid sequence FP-CATH of the mature peptide of Deinagkistrodon acutus obtained, the FP-CATH9 sequence is obtained, and its full sequence is synthesized by an automatic peptide synthesizer and desalted and purified by HPLC reversed-phase column chromatography; its purity is identified by the HPLC method of high-performance liquid chromatography, the sequence is identified by LC-MS / MS, the isoelectric point is measured by isoelectric focusing electrophoresis, the secondary structure is measured by circular dichroism spectroscopy, and the tertiary structure of the amino acid sequence is predicted by SWISS software.

3. The application of the antibiotic adjuvant according to claim 1, characterized in that the antibiotic adjuvant is used in combination with minocycline or tigecycline to prepare an antibacterial drug or composition.

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