Discovery Of Active Molecules Against Drug-resistant Bacteria From Dendrobium Huoshanense And Cytospora Chrysosperma

Clinically resistant bacteria refer to pathogenic bacteria that have developed resistance to traditional antibacterial drugs,which is a self-evolution result of bacteria’s long-term struggle with antibacterial drugs,as well as the"special ability"of bacteria to survive themselves.After Fleming discovered penicillin in the 1920s,the battle between humans and bacteria kicks off.Then,with the new antibiotics are discovered and abused,the vast majority of sensitive bacteria were eliminated,the relatively insensitive bacteria developed drug resistance through continuous reproduction and evolution,and eventually replaced the sensitive strains.As early as 2014,WHO（World Health Organization）published an article saying that clinically drug-resistant bacteria are spreading worldwide and gradually becoming a global public problem.recent years,many experts have done systematic research on this issue,they are looking for new antibiotics from animals and plants,microorganisms and chemical synthesis.Our research group has long been committed to discovering novel lead compounds with anti-clinical drug-resistant bacteria from microorganisms and traditional Chinese medicine.In the previous activity screening experiments,we found that the fermentation product of HYQZ-931（Cytospora chrysosperma）,a plant endophytic fungus of Hippophae rhamnoides,and the alcohol extract of Dendrobium huohanense displayed inhibitory activity against multiple clinically resistant bacteria.So,we selected them as the research target.First,Dendrobium huohanense was extracted with anhydrous ethanol to obtain its crude extract,and large-scale fermentation and culture of Cytospora chrysosperma was carried out to extract its secondary metabolite,then through various separation and purification methods to separate and purify its chemical components.The structure of each monomer compound was determined according to the theory and method of modern spectroscopy and related literature.Finally,activity screening was performed.This paper consists of four chapters.In the first chapter,the study of Dendrobium nobile Lindl is reviewed based on the structure of chemical constituents,including sesquiterpenes,phenanthrenes,bibenzyl,alkaloids,polyketones,phenolic acids and coumarins,by referring to relevant literature.In chapter 2,we mainly discussed the chemical composition and biological activity of Dendrobium huoshanense.In the early stage,the fresh Dendrobium Huoshanense collected in Huoshan County,Anhui Province,was crushed and impregnated with a certain amount of ethanol for three times.Combine the extracts and concentrate under reduced pressure to obtain its first extract.Then washed with water,extracted with ethyl acetate and ultimately obtain its crude extract,and isolated 33 compounds from the crude extract through a series of purification experiments,including 1 novle benzyl compounds and 1 novel Phenylpropanoid compounds;their structures were determined by modern spectroscopic methods(¹H NMR、¹³C NMR、HSQC、HMBC、¹H-¹HCOSY,NOESY、HRESIMS、UV、IR、ECD,optical rotation,X-ray crystal diffraction,etc.)combined with relevant literature,simultaneously tested their biological activity against multiple strains of clinically resistant bacteria and human tumor cells,also compounds3 and 4 were successfully found to exhibit potent antibacterial activity against methicillin-resistant Staphylococcus aureus（MIC,1.56μg/m L）.In chapter 3,we mainly discussed the chemical composition and biological activity of Cytospora chrysosperma.Firstly,PDB medium was used to produce the seed liquid of Cytospora chrysosperma,and then rice+MEB solid medium was used to conduct large-scale solid fermentation culture of Cytospora chrysosperma.After fermentation,the crude extract was extracted by methanol,subsequently dissolved in water,extracted with ethyl acetate,and oncentrated under reduced pressure to obtain the crude extract of Cytospora chrysosperma,ultimately obtained 13 compounds through various chromatographic separation methods,Including 3 novel cytochalasins and 1 novel cyclobutane dimer compound.Their chemical structures were determined by modern spectroscopic methods combined with relevant literatures,and the bioactivity against multiple clinically resistant bacteria,crop pathogenic fungi and human tumor cells was also tested,finally,the novel cytochalasins 1 and 3 were successfully found that have certain antibacterial activity against methicillin-resistant Staphylococcus aureus and multidrug-resistant Enterococcus faecium（MIC,25μg/m L）.at the same time,based on the OSMAC strategy,we further explored the biosynthetic potential of Cytospora chrysosperma.The fourth chapter mainly summarizes and looks forward to the whole thesis.