Tandem mass spectrometry applied in the development of analytical methods in identification of polar microbial metabolites

• Grant number: 21/13532-7
• Support Opportunities: Regular Research Grants
• Start date: March 01, 2022
• End date: August 31, 2024
• Field of knowledge: Physical Sciences and Mathematics - Chemistry - Organic Chemistry
• Principal Investigator: Luiz Alberto Beraldo de Moraes
• Grantee: Luiz Alberto Beraldo de Moraes
• Host Institution: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

The importance of microbial natural product chemistry as a source of bioactive molecules is undoubted. The isolation and identification of secondary metabolites produced by microorganisms is very important for the chemical, pharmaceutical and agrochemical industries. Studies have revealed that some microbial secondary metabolites, encoded in the genomes, remain to be discovered, probably because these genes are not transcribed under laboratory conditions. Several approaches have been taken in an attempt to activate these cryptic biosynthetic pathways, which include modifications of the culture media (OSMAC-, one strain of many compounds), co-cultivation, mixed cultivation, among others. Likewise, different analytical methodologies based on LC-MS/MS, such as dereplication, molecular networking and MS-imaging, have been developed in the last decade with the aim of accelerating the annotation or identification of secondary metabolites present in crude extracts. However, few studies are described in the literature aiming at the development of sample preparation methodologies for the isolation of polar metabolites produced by microorganisms. For this reason, this research project aims to evaluate different sample preparation techniques for the identification of bioactive polar secondary metabolites. For this, new sample preparation methodologies will be evaluated using SPME in the Coated Blade Spray (CBS) form, which allows a direct coupling to the MS ionization source. CBS is one of the most comprehensive technologies in sample preparation. Liquid-liquid extraction techniques in conventional mode and by "QuEChERs" will also be evaluated. For the development of this methodology, commercial polar metabolites will be used, such as amino glycosides, phosphonates and sulphonates. The developed methodology will be applied in samples from fermentation processes with actinobacteria producing amino glycosides and in the collection of microorganisms present in our laboratory. (AU)