CN115350198B - Application of epilithocholic acid - Google Patents

Application of epilithocholic acid Download PDF

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Publication number
CN115350198B
CN115350198B CN202210961982.5A CN202210961982A CN115350198B CN 115350198 B CN115350198 B CN 115350198B CN 202210961982 A CN202210961982 A CN 202210961982A CN 115350198 B CN115350198 B CN 115350198B
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Prior art keywords
acid
epilithocholic
streptococcus mitis
application
test
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CN202210961982.5A
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Chinese (zh)
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CN115350198A (en
Inventor
王帅
王玉桂
郭爱疆
邹扬
张少华
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Lanzhou Veterinary Research Institute of CAAS
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Lanzhou Veterinary Research Institute of CAAS
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00Medicinal preparations containing organic active ingredients
    • A61K31/56Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • A61K31/575Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of three or more carbon atoms, e.g. cholane, cholestane, ergosterol, sitosterol
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/02Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00Drugs for disorders of the respiratory system
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00Drugs for disorders of the urinary system
    • A61P13/08Drugs for disorders of the urinary system of the prostate
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00Drugs for dermatological disorders
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00Drugs for disorders of the senses
    • A61P27/16Otologicals
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04Antibacterial agents
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61PSPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00Drugs for disorders of the cardiovascular system
    • YGENERAL 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
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02ATECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention discloses an application of a known compound, namely, epilithocholic acid, in preparing medicines for resisting streptococcus mitis ATCC 49456. The experiment of the invention shows that the epilithocholic acid has good activity of inhibiting streptococcus mitis, and is a potential drug capable of replacing the existing antibiotic to inhibit streptococcus mitis.

Description

Application of epilithocholic acid
Technical Field
The present invention relates to the use of an epilithocholic acid (Epialolishocholic acid), a known compound.
Background
Bile acid is an active ingredient in bile, and is one of important ingredients involved in the digestive function of bile. The content change has important reference value in liver disease diagnosis, and the published literature discloses that bile acid has the effect of inhibiting tumors. In addition, chinese patent application 202111019726.6 discloses that cholelithiaacid has antibacterial activity against staphylococcus aureus MRSA and MSSA, escherichia coli esbls+, escherichia coli Esbls-, and pseudomonas aeruginosa. The epilithocholic acid is one of the most important secondary bile acids in intestinal tracts, and the molecular formula is as follows: c (C) 24 H 40 O 3 The structural formula is shown as follows:
disclosure of Invention
The invention discloses application of table allophanate.
The application of the epilithocholic acid in preparing the medicine for resisting streptococcus mitis is provided.
The streptococcus mitis belongs to alpha-hemolytic streptococcus (also called alpha-hemolytic streptococcus), is one of normal flora of parts of the oral cavity, the digestive tract, the genital tract and the like of a human body, is a conditional pathogenic bacterium, can cause toxic shock syndrome, subacute endocarditis, prostatitis and the like, and can also cause pneumonia, pericarditis, peritonitis, salivary gland inflammation, orofacial infection, odontogenic infection, otitis media, sinusitis and the like. It has been reported that this pathogen was isolated in the body of the affected pigs and resulted in severe morbidity and mortality in piglets, suggesting that streptococcus mitis is also pathogenic to pigs. Among the common antibiotics for pediatrics, the streptococcus mitis has different degrees of drug resistance to aztreonam, ceftriaxone, piperacillin/tazobactam and amoxicillin/clavulanate potassium, and the drug resistance rate reaches 42.3% -48.1%. These antibiotics are still currently the usual drugs for pediatric pneumonia. Bacterial resistance is a global problem, but the development of corresponding antibiotics is slow and the form is very severe, so a new antibacterial drug or antibacterial product is urgently needed to inhibit bacteria. The invention is a technical proposal for solving the defects of the prior art, and is a potential drug capable of replacing the prior antibiotics to inhibit streptococcus mitis.
Experiments show that the epilithocholic acid has good activity of inhibiting streptococcus mitis.
Drawings
Fig. 1: and (3) a graph of the relationship between the concentration of the epilithocholic acid and the growth inhibition rate of streptococcus mitis.
Fig. 2: a graph of chloramphenicol concentration versus growth inhibition rate of streptococcus mitis.
Detailed Description
The invention is illustrated below with reference to examples, in which an in vitro antibacterial activity test was performed on lithocholic acid using a trace broth double dilution method, and the antibacterial activity against streptococcus mitis was examined, and experimental system evaluation was performed on chloramphenicol as a control.
Test article and control
1.1 test article
Name: and (3) table lithocholic acid, which is commercially available.
Before the test sample is used, ethanol is used for preparing the test sample into 5mg/mL of maximum concentration, and the test sample is diluted in double in sequence.
1.2 control
Name: chloramphenicol, commercially available.
(II) test Strain and method for culturing the same
2.1 Strain: streptococcus mitis ATCC49456
2.2 sources of strain: purchased from Beijing Bai Ou Bo Wei Biotechnology Co.
2.3 culture conditions: todd-HewittBroth, incubation at 37℃for 24h-48h.
2.4 starting seed and culturing the strain
The outside of the penicillin bottle is wiped by 75% alcohol cotton, and a plastic cover and an aluminum cover are removed by using a nipper pliers in a safety cabinet. Slowly opening the rubber plug, sterilizing the bottle mouth part with 75% alcohol cotton, injecting 0.5ml liquid culture medium re-dissolving freeze-dried powder with a sterile straw, inoculating the suspension on the culture medium, culturing at 37 ℃ for 24 hours, and then picking single bacterial colony to culture and increase bacteria in the liquid culture medium under the same culture condition.
2.5 preparation of working solution of bacterial liquid
When the OD600 value reaches about 0.5 (about 10 9 CFU/ml), subjecting to gradient dilution (0, 10, 100, 1000, 10000, 100000, 10000 times) with THB broth, inoculating 100 μl of bacterial suspension onto solid culture medium, culturing for 24 hr, counting viable bacteria, calculating bacterial suspension concentration, and diluting the bacterial suspension to obtain bacterial suspension final concentration of about 2×10 6 CFU/ml. The strain operation is carried out under the aseptic condition, and after the experiment is finished, all consumable materials are sterilized and then discarded.
(III) determination of the minimum inhibitory concentration required to inhibit 50% of the test bacteria (MIC 50)
3.1 test method: MIC50 determination was performed by a micro broth dilution method.
The experiments were divided into positive control (i.e. bacterial suspension without drug), negative control (medium with vehicle ethanol) and experimental (bacterial suspension with different concentrations of lithocholic acid). Each of the above groups was provided with 3 parallel groups. The experiment solution of the epilithocholic acid with the concentration of 100 mug/ml is diluted by times in a liquid culture medium to obtain the final concentration of 100 mug/ml, 50 mug/ml, 25 mug/ml, 12.5 mug/ml, 6.25 mug/ml, 3.125 mug/ml and 0 mug/ml. The concentration is 2X 10 6 CFU·mL -1 The bacterial suspension and each group of liquid medicine are respectively inoculated into a sterile 96-well plate according to the proportion of 1:1 and 100 mu L, so that the final concentration of the bacterial suspension is 1 multiplied by 10 6 CFU/ml (containing 10) 5 CFU/well), and cultured under appropriate culture conditions for 20 hours. Meanwhile, chloramphenicol as a control was set so that its final concentration was 32. Mu.g/ml, 16. Mu.g/ml, 8. Mu.g/ml, 4. Mu.g/ml, 2. Mu.g/ml, 1. Mu.g/ml, and 0. Mu.g/ml, respectively. After the culture is finished, visual observation is carried out under a black background, and the lowest drug concentration without bacterial growth in the plate is the Minimum Inhibitory Concentration (MIC) of the lithocholic acid. MIC determination experiments for all species were repeated 3 times. The method for calculating the OD600 value, the test samples with different concentrations and the control bacteriostasis rate of the suspended bacteria comprises the following steps: antibacterial ratio (%) =1- (test OD value-negative control OD value)/(positive control OD value-negative control OD value) ×100%. The MIC50 is calculated by curve fitting a function.
3.2 judgment of results
The results of this test are shown in Table 1, and the inhibition rates are shown in FIG. 1. The test is only meaningful when the bacteria grow significantly in the positive control wells (i.e. without drug). The results showed that bacteria grew normally in the positive control wells (i.e., without drug) and that the vehicle control medium wells grew aseptically. The MIC value of the control chloramphenicol for Streptococcus mitis was 4. Mu.g/ml, the results are shown in Table 2, and the inhibition ratio is shown in FIG. 2, which meets the "antimicrobial susceptibility test execution Standard" of the American clinical laboratory standardization Committee (National committee for clinical library standardization, NCCLS) 2001: the chloramphenicol in the eleventh edition of information journal M100-S11 is less than or equal to 4 mug/ml for streptococcus mitis, which shows that the experimental method is reliable.
Statistical results show that: the MIC50 of epilithocholic acid for the tested Streptococcus mitis was 6.526 μg/ml. In conclusion, the epilithocholic acid has good activity of inhibiting streptococcus mitis.
TABLE 1 absorbance measurements of the effect of different concentrations of epilithocholic acid on growth of Streptococcus mitis
Table 2: absorbance measurement of the effect of different chloramphenicol concentrations on growth of Streptococcus mitis

Claims (1)

1. Use of epilithocholic acid in the preparation of a medicament against streptococcus mitis ATCC 49456.
CN202210961982.5A 2022-08-11 2022-08-11 Application of epilithocholic acid Active CN115350198B (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112245423A (en) * 2020-10-21 2021-01-22 兰州大学 New application of acylated homoserine lactone compound in treatment of oral diseases
CN113559013A (en) * 2021-09-01 2021-10-29 南充市中心医院 Application of lithocholic acid compound in preparation of antibacterial product

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11077123B2 (en) * 2016-02-29 2021-08-03 Tel Hashomer Medical Research Infrastructure And Services Ltd. Use of bile acids and bile salts as anti bacterial agents for inhibition of bacterial conjugation and horizontal gene transfer

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112245423A (en) * 2020-10-21 2021-01-22 兰州大学 New application of acylated homoserine lactone compound in treatment of oral diseases
CN113559013A (en) * 2021-09-01 2021-10-29 南充市中心医院 Application of lithocholic acid compound in preparation of antibacterial product

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
Human gut bacteria produce ΤΗ17-modulating bile acid metabolites;Donggi Paik等;《Nature》;第603卷;第907-938页 *
The Role of Bile Acid in Improving Glucose Tolerance of Non-Obese Diabetic Rats After Proximal Small Bowel Bypass;Zhihua Zheng等;《Frontiers in Physiology》;第13卷;第1-10页 *

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