Metabolome Variation Of Mariannaea Pruinosa At Different Culture Time

Mariannaea pruinosa Liang sp.nov.is the anamorph of Cordyceps pruinosa Petch.Cordyceps pruinosa and its anamorph have been reported possessing multi-bioactivities such as immuno regulation,antibactierium,antifungus,antitumour and anti-irradiation.To figure out metabonomic discipline of Mariannaea pruinosa cultured with different time we employed an HPLC-MS based metabonomics method to study the metabonomics variations of Mariannaea pruinosa during different cultural time.The result can provide scientific reference for fermentation optimization and metabolites uitilization of the fungus,meanwhile it can also be theoretical basis for the fungal metabolotes regulation and products developmentFirst of all,we detected all small molecular metabolites in the fermentation broth of Mariannaea pruinosa at different culture time.Then,multivariate statistics based metabonomic methods were used to analyse the detected data.PCA(Principal components analysis)showed that metabolites from fermentation broth of different time cultured Mariannaea pruinosa can be divided into three stages: prophase(3-9d),metaphase(12-15d)and anaphase(20d).Based on the putatively identified metabolites and analysis of OPLS-DA(Orthogonal projection on latent structure-discriminant analysis),it was found that the fungus at prophase synthesized more of sitosterol,AIR,farinosone A,monoterpene glycoside,xanthosine monophosphate,butanoic acid,gamma-amino butyric acid,ketone xanthones,geniposide,altersolanol A,cordyformamide,protoemetine,phosphorus ester acyl serine and choline etc.At metaphase we detected more of CDP-glycerol,UDP-N-acetyl-alpha-Dglucosamine,oleandolide,citric acid,indole pyruvic acid,leucine,p-coumaroyl putrescine,phosphoricacid,hydromorphone,guanosine,sphingosine,phytosphingosine,S-adenosyl-homocysteine,UDP-rhamnose,UDP-N-acetyl-D-mannosamine,4-hydroxyl mandelic acid ethylester and 4-hydroxyazelaic acid etc.which related to mycelial growth and spores reproduction,energy production and immune defense.Some of the metabolites are intermediates.At anaphase the fungus metabolites more of 28,30-dinor-17,19,21-gammaceratriene,4-methyl-2-oxo pentanoate,militarinone B,cordyol A,malate,citric acid,leucine,nicotinic acid(vitamin PP),flavin nucleotide(FMNH2),zygosporin D,CTP,2-phosphoglycerate,phosphoenol pyruvate phosphoenolpyruvate,PEP,mannitol and uracil.Some of the compounds are relation to sporulation,oxidative stress,anti-bacteria,anti-inflammatory and immunity enhancement.Of all above metabolites cordyformamide,farinosone A,prosafrinine,militarinone B,cordyol A,phytosphingosine,mannitol and zygosporin etc.have strong bioactivity and utilization potencial.The antibiotics prosafrinine was originaly isolatied from a plant,but our study revealed that it can be produced by M.pruinosa,therefore can be automaticly produced by fermentation in factory.?-sitosterol has acivities such as cholesterol lowering,antitumour,anti-inflammatory,anti-tussive,oxygen free radicals scavenging and antioxidation.This study revealed that sitosterol reached maximum vlevel after 6 days fermentation of the fugus,therefore this compound can be havested at the fermentation prophase.Mannitol in the fermentation broth reached the highest level at 15 days fermentation therefore this compound should be better havested at 15 days fermentation.This study revealed for the first time the metabolic relationships between GABA,phytosphingosine and sphingosine.GABA is a important regulatary factor in bacteria,plant and animals,meanwhile,sphingosines play also very important role in cell signal transportation and immune regulation.Because of similar structures phytosphingosine and sphingosin can be possiblly interconversed,or come from the same precursor,but the relationship them and GABA need further study.More impotantly,we found the metabolism of cordycepoid A was related to energy production.It is possible that the pigment can absorb luminous energy and produce CTP for energy production for the fungus.This is worth of further study.During the metaphase and anaphase of fermentation the production of nucleoside phosphates were significantly increased.The reason is possible that through glycosylation the fungus can detoxify a unmber of toxic secondary metabolites.