Studies Of Biofllm Formation And Drug Interference Mechanisms Of Candida Albicans By Combination Of Untargeted And Targeted Metabolomics Modes

Candida albicans (C. albicans) is the most common fungal pathogen which can attach toinert and biological surfaces to form biofilm. C. albicans infection has been a clinical thornychallenge since it can intrinsically resistant to most antifungal agents and host defensesthrough various resistance mechanism.In recent years, with the development of metabolomics techniques, metabolites ofdifferent forms of fungal were characterized and successfully apply to metabolic profilinganalysis. Our previous research displayed that amino acids metabolism, as an important partof the regulation of metabolic pathways, played a key role in the process of C. albicansmorphological changes. On that basis, LC-MS and GC-MS were applied to targeted anduntargeted metabolomics study. Combined with metabolomics data mining andpharmacological effects verification, C. albicans morphological changes as well as traditionalChinese medicine antifungal pharmacodynamics mechanism were discussed for the purposeof offering approach reference to fungal metabolic research.In response to the polarity and mass spectrometry characteristics of amino acids, anIP-PGC-TOF/MS system was developed. On the basis of the matrix effect, precision,repeatability and recovery evaluation, three isotope labeled amino acids were chosen asinternal standard, and a quantitative analysis method for22underivatized amino aicds in C.albicans cells was developed. Both of the lowest limit of detection and limit of quantificationachieve nanogram. The established method was an offer of assistance for the targeted anduntargeted profiling of C. albicans.A combination of metabolic fingerpriting analysis and metabolic target analysis wereapplied to the study of the mechnism of C. albicans biofilm formation. On the basis ofuntarged profiling metabolism,16differential metabolites were identified as potentialbiomarkers. A quantitative analysis by IP-PGC-TOF/MS system was followed, and the levelof amino acid metabolism changes was carefully characterized, a series of potentialbiomarkers related to biofilm formation was identifeid and found mainly involed in ornithineand lysine related metabolism. Ornithine was obseved down-regulated obviously in biofilmcells. Further experiments were carried out using ornithine decarboxylase-nagative (spe1Δ)mutant, cell surface hydrophobicity assay (CSH) verified the key role of ornithine metabolism in biofilm formation.A metabolomic approach based on targeted and untarged profiling has been applied tocharacterize the metabolic profile associated with baicalein treatment and investigate theintervention mechanism of baicalein. Based on both GC-MS and IP-PGC-TOF/MS assay,20and21potential biomarkers have been screened out respectively as being primarily involvedin glycolysis, ocidative stress response, amino acids metabolism, citrate cycle, phospholipidand sphingolipid catabolism. In addition, quantitative analysis of22amino acids was alsocompleted. Thus demonstrated that baicalein could probably treated C.albicans by disturbingenergy metabolism, cell membrane function as well as quorum sensing synthetize. With thealtered metabolism pathways as possible drug targets, baicalein played the role of antifungal.The established targeted and untargeted profiling metabolism approach were successfullyapplied to the research of C. albicans. Compared with traditional strategy, our system couldprovide more sensitive metabolites identification ability as well as more accuratequantification of amino acids, thus provided new ideas for the metabolism research of C.albicans biofilm and the development of new antifngal agents.