New chromatographic strategies for metabolomics and exogenous compounds analysis in microorganisms and food matrices

Author(s)

João Raul Belinato de Souza

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Química
• Defense date: 2020-11-11

Examining board members

Fabio Augusto; Carla Beatriz Grespan Bottoli; Gisele André Baptista Canuto; Marcio Pozzobon Pedroso; Taicia Pacheco Fill

• Advisor: Fabio Augusto; Katia Cristina Kupper

Abstract

Metabolomics has been rapidly developed in recent years due to major advances in analytical instrumentation, particularly chromatographic and mass spectrometry methods. It has also been supported by new and more efficient computational tools. In addition, the use of more robust extraction techniques has supported the increasingly detailed assessment of highly complex systems. Thus, this work aimed to use some of these analytical approaches based on metabolomic strategies and applied them to problems involving biological control using microorganisms, early detection of fungal diseases in food and quantification of exogenous compounds in food matrix. The first application explored the use of yeasts as biological control agents against different post-harvest citrus phytopathogens. The use of untargeted approaches and sophisticated analytical strategies such as GC×GC and UHPLC-MS/MS allowed a more detailed understanding of the metabolites involved in interactions between microorganisms responsible for the antagonistic action. Moreover, the developed strategy allows to surmise the potential anti-phytopathogenic action of these yeasts, being a great advance in biological control¿s field. Then, using a targeted approach, a known metabolite pattern of A. niger was studied aiming to detect the fungus with only one day of growth. The approach was later extended to the analysis of contaminated grapes, which were analyzed using an in situ-HS-SPME and GC×GC-ToF-MS strategy assisted by multivariate analysis. The results showed that this set of metabolites can be used to confirm the presence of the pathogen in vitro and in two varieties of table grapes after only one day of contamination. Finally, a targeted approach was used in the determination of exogenous compounds, such as pesticides, in honey. In this case, a fully automated quantitative method for the determination of pyrethroids in honey was developed using overcoated SPME fibers compatible with the matrix and GC-MS. The proposed method has been optimized using multivariate tools and its validation has provided adequate figures of merit. Thus, the potential of this method, as well as the overcoated fibers prospects are very promising. Especially as there are several obstacles associated with challenging matrices and minimal sample handling through automation. Therefore, the strategies implemented here have significantly contributed to expand the understanding of these metabolomic systems, as well as the application of new analytical approaches that sustain the growth of this area and encourage new discoveries that benefit society (AU)

• FAPESP's process: 16/20547-2 - Characterization of the metabolome of anti-phytopatogen microorganisms by Multidimensional Gas Chromatography and Simple / Tandem Mass Spectrometry (GC×GC-QMS and GC×GC-MS/MS)
• Grantee: Joao Raul Belinato de Souza
• Support Opportunities: Scholarships in Brazil - Doctorate