Activity Tracking Of Anti-mycobacterial Components Of Thistle

Tuberculosis(TB)is an infectious disease caused by Mycobacterium tuberculosis(MTB).It can invade all organs throughout the body,but pulmonary is most common and it is still one of the most fatal infectious diseases in the world.In recent years,the number of patients suffering from tuberculosis in the world has been declining at a slow rate,but the number of multidrug-resistant tuberculosis(MDR-TB)and extensively drug-resistant tuberculosis(XDR-TB)is increasing,which makes tuberculosis treatment face severe challenges.Therefore,the discovery of new anti-tuberculosis drug targets and the development of new anti-tuberculosis drugs have become an urgent problem to be solved.In this paper,studying on the active component against Mycobacterium tuberculosis from the roots of Cirsium japonicum DC.and exploring its anti-mycobacterium tuberculosis mechanism.The ethanol extract of Cirsium japonicum DC.was obtained by ethanol refluxing method.The ethanol extract of Cirsium japonicum DC.was carried out to obtain the extraction part of the petroleum ether layer,the extraction part of the ethyl acetate layer,and the remaining layer.Using bioassay-guided isolation method,it was found that the active site was located in the ethyl acetate part.The ethyl acetate part was separated by silica gel column chromatography,and putting the same components together after TLC detection.After repeated silica gel column chromatography,the monomer compound HP01 inhibiting mycobacterium tuberculosis was obtained.Its MIC of anti-mycobacterium tuberculosis is 160 ?g/m L by the activity test.Compound HP01 is yellow powder.According to literature spectral data,it was identified as quercetin.It was found that the monomer compound HP01 had an additive effect with pyrazinamide,but had no antagonistic effect with isoniazid and rifampicin by the checkerboard test.After the monomer compound HP01 forms a metal complex with copper ions,its activity increases.In order to explore the anti-mycobacterium tuberculosis mechanism of the monomer compound HP01,molecular docking technology was used to make docking between the monomer compound HP01 and the Kat G enzyme,the result of molecular docking showed that the monomer compound HP01 had a strong affinity with Kat G enzyme.Scanning electron microscope was used to observe the morphology of Mycobacterium tuberculosis,scanning electron microscopy images showed that the monomer compound HP01 could destroy the cell wall of Mycobacterium tuberculosis.By using ultraviolet spectrophotometry to test the Kat G enzyme activity,the result showed that the monomer compound HP01 had an inhibitory effect on the Kat G enzyme of Mycobacterium tuberculosis.The preliminary results speculate that Kat G enzyme is the main target of the monomer compound HP01.Real-time fluorescent quantitative PCR technology(q RT-PCR)was used to study the relative transcription of Kat G gene after the monomer compound HP01 interfered with Mycobacterium tuberculosis.The experimental results showed that the transcription of Kat G gene was significantly compensatory up-regulated after Mycobacterium tuberculosis was intervened with the monomer compound HP01,which indicates the monomer compound HP01 may be the Kat G enzyme inhibitor of Mycobacterium tuberculosis.In summary,quercetin is a molecule with anti-mycobacterium tuberculosis activity from the root of Cirsium japonicum DC.,and it produces antibacterial effect by inhibiting Kat G enzyme.The thesis lays the foundation for the development of anti-tuberculosis drugs with a new mechanism of action.