Furocoumarin compound and application thereof in detection of mycobacterium tuberculosis

A technology of mycobacterium tuberculosis and compounds, applied in organic chemistry, sugar derivatives, analytical materials, etc., can solve problems such as low signal-to-noise ratio and instability

Active Publication Date: 2021-06-18
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, the CDG-DNB3 developed by Rao et al. at Stanford University can rapidly and specifically image Mycobacterium tuberculosis, but the molecule has a low signal-to-noise ratio and is unstable under clinical detection conditions (alkaline)

Method used

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  • Furocoumarin compound and application thereof in detection of mycobacterium tuberculosis
  • Furocoumarin compound and application thereof in detection of mycobacterium tuberculosis
  • Furocoumarin compound and application thereof in detection of mycobacterium tuberculosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] Embodiment 1 The fluorescence response of the compound of the present invention to Rv2466c enzyme

[0085] Instrument: multifunctional microplate reader ( Flash, Thermo Scientific).

[0086] Reagents and materials: compound (A1-A6), Rv2466c, MSH, Tris-Cl (pH 7.5), DMSO, 96-well microtiter plate (Corning, 3603).

[0087] Operation: 1) Prepare 50mM Tris-Cl (pH 7.5) buffer solution containing 0.5mM MSH, and put 20mL buffer solution into two sterile tubes. 2) Add a certain amount of Rv2466c to one of the tubes so that the final concentration is 200 μg / mL, which is the enzyme-containing working solution, and add the same amount of Tris-Cl buffer solution to the other tube as the enzyme-free control solution. 3) The compounds of the present invention were respectively dissolved in DMSO to prepare a solution of 25 μg / mL, and 20 μL of each compound was added to four wells of a 96-well plate, two of which were used as a control group without enzymes , and the other two wells...

Embodiment 2

[0089] Embodiment 2 The specificity experiment of compound A6 of the present invention

[0090] Instrument: Fluorescence microscope (Delta Vision high-resolution live cell imaging system).

[0091] Reagents and materials: compound A6, intermediate 12, Bacillus subtilis (Bs), Escherichia coli (Ec), Enterococcus faecalis (Ef), Listeria monocytogenes (Lm), Staphylococcusaureus (Sa), Mycobacterium smegmatis (M.smeg), cultured in LB base, THY medium, 7H9 medium.

[0092] Operation: Prepare the compound A6 and intermediate 12 at a concentration of 10 mM. After the bacteria are cultured to the logarithmic growth phase, take 200 μL of bacterial liquid and add 2 μL of bacterial liquid to 2 μL A6 and intermediate 12 respectively. After incubating at 37°C for 30 minutes, prepare a slide sample and examine it under a fluorescence microscope. observe. The results show that the compound of the present invention has specificity to mycobacteria, and does not stain to other Gram-negative bac...

Embodiment 3

[0094] Embodiment 3 The dyeing ability of compound A6 of the present invention to Mtb

[0095] Instrument: fluorescence microscope.

[0096] Reagents and materials: Compounds A1 and A6, Mycobacterium tuberculosis (Mtb), 7H9 medium.

[0097] Operation: Prepare the compounds A1 and A6 at a concentration of 10 mM respectively, culture the bacteria to the logarithmic growth phase, take 200 μL of the bacterial liquid and add ebselen (Eb, the final concentration is 50 μg / mL), incubate for 3 hours and centrifuge to remove ebselen, respectively. Add 200 μL 7H9 medium and 2 μL A1, A6, and incubate at 37°C for 30 minutes, prepare slide samples and observe under a fluorescence microscope. The results are as follows: Figure 5 .

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Abstract

The invention discloses a furocoumarin compound capable of labeling single cells of mycobacterium tuberculosis, the compound has a structure as shown in a formula I, a formula II or a formula III, and the compound can realize effective and real-time detection of clinical suspected tuberculosis patients through fluoroscope.

Description

technical field [0001] The invention relates to the technical field of mycobacterium tuberculosis detection, in particular to furanocoumarin compounds and their application in the detection of mycobacterium tuberculosis. Background technique [0002] Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is one of the deadliest infectious diseases in the world. According to WHO statistics, about 4,000 people die of tuberculosis every day in the world, and 30,000 people are infected with Mycobacterium tuberculosis. Currently, about 1 / 3 of the world's population is infected with Mtb, and 90% of them are asymptomatic latent tuberculosis (latentTB, LTB) infected. Mtb is mainly transmitted through air droplets. In recent years, there have been more and more drug-resistant tuberculosis, and even all-drug-resistant strains have emerged, which has brought great challenges to the prevention and treatment of TB. [0003] Achieving early diagnosis of tuberculosis and carrying...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H19/056C07H5/06C09K11/06G01N21/64
CPCC07H19/056C07H5/06C09K11/06G01N21/6402C09K2211/1088C09K2211/1059C09K2211/1029Y02A50/30
Inventor 刘刚李雪媛耿鹏飞洪小巧
Owner TSINGHUA UNIV
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