Identification of Fusarium oxysporum Fungus in Wheat Based on Chemical Markers and Qualitative GC-MS Test

Carlos Senes,Willian Costa,Terezinha Svidzinski,Cláudio Oliveira,Nayara Saldan

doi:10.21577/0103-5053.20180143

Abstract

Fusarium oxysporum is usually found in foods such as corn, wheat, soybeans and barley, and can cause economic losses and risks to health in humans and animals due to the production of mycotoxins. The conventional microbiological test for fungal identification is based on morphological characteristics, requires specialists, and it is very time-consuming as it is necessary to carry out a culture of the fungus for two weeks before microorganism identification. Nowadays, researchers have made efforts to develop alternative analytical methods with advantages such as shorter time of analysis and better accuracy and reproducibility when compared to microbiological methods. In this way, in the present work, a qualitative analytical method to detect the secondary metabolites of Fusarium oxysporum in culture media by gas chromatography coupled to mass spectrometry (GC-MS) was developed with the intention of relating the presence of these chemical compounds to contamination by the fungus. The method comprised liquid-liquid or liquid-solid extractions with ethyl acetate from liquid and solid culture media, respectively, followed by GC-MS analysis, which was able to identify the presence of fusaric acid, 13-socosenamide, 5-(4-hydroxybutyl)-2-pyridinecarboxylic acid and toxin HT-2. The presence of these compounds was confirmed by high-performance liquid chromatography coupled to mass spectrometry quadrupole time of flight (HPLC-MS/QTOF).

Highlights

Fungi are microorganisms that present chromosomes and nucleoli enclosed in a cell membrane, being classified as eukaryotic
The objective of the present work is to develop a qualitative methodology using gas chromatography coupled to mass spectrometry (GC-MS) to separate and identify chemical substances produced by F. oxysporum in traditional culture media and in wheat samples that can be used as chemical markers of the fungus, opening the possibility to develop an analytical method to identify the fungus chemically with greater speed and sensitivity
The mass spectrum of the peak with retention time of 34.96 min was attributed to the mycotoxin fusaric acid (m/z 179), as the ion of m/z 164 indicated the loss of methyl fragment and the ion fragment of m/z 136 indicated the loss of C2H4.5,6,21 The mass spectrum of the peak with retention time of 36.33 min was attributed to 5-(4-hydroxybutyl)2‐pyridinecarboxylic acid (m/z 195), which is a derivative of fusaric acid, and the ions of m/z 164 and 122 indicated the loss of CH3O and C4H9O fragments, respectively

Summary

Introduction

Fungi are microorganisms that present chromosomes and nucleoli enclosed in a cell membrane, being classified as eukaryotic. They can be unicellular or multicellular, and are classified as heterotrophic. Reproduction usually occurs through spores and can be sexual or asexual.[1] Organic matter and humidity are important for their growth, but they can survive in dry environments through the production of aridity-resistant spores.[1]. Toxigenic fungi produce toxic secondary metabolites (mycotoxins) during their life, which can be carcinogenic in human and animals.[2] Fungi can enter the food chain through food contamination, which occurs by the contact with spores present in the environment, in the soil, or during harvesting, storage, and transport,[3] and

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Conclusion