Multi-capillary column-ion mobility spectrometry of volatile metabolites emitted by Saccharomyces cerevisiae

Abstract

Volatile organic compounds (VOCs) produced during microbial fermentations determine the flavor of fermented food and are of interest for the production of fragrances or food additives. However, the microbial synthesis of these compounds from simple carbon sources has not been well investigated so far. Here, we analyzed the headspace over glucose minimal salt medium cultures of Saccharomyces cerevisiae using multi-capillary column-ion mobility spectrometry (MCC-IMS). The high sensitivity and fast data acquisition of the MCC-IMS enabled online analysis of the fermentation off-gas and 19 specific signals were determined. To four of these volatile compounds, we could assign the metabolites ethanol, 2-pentanone, isobutyric acid, and 2,3-hexanedione by MCC-IMS measurements of pure standards and cross validation with thermal desorption-gas chromatography-mass spectrometry measurements. Despite the huge biochemical knowledge of the biochemistry of the model organism S. cerevisiae, only the biosynthetic pathways for ethanol and isobutyric acid are fully understood, demonstrating the considerable lack of research of volatile metabolites. As monitoring of VOCs produced during microbial fermentations can give valuable insight into the metabolic state of the organism, fast and non-invasive MCC-IMS analyses provide valuable data for process control.

Figure 1

(A) Experimental set-up for the on-line MCC-IMS measurements of fermenter off-gas. (B) Working principle of the ion mobility spectrometer; adapted from [33].

Figure 2

MCC-IMS topographic plot of sterile Verduyn medium. The reaction ion peak (1/K₀ = 0.5 Vs cm⁻²) was compensated by the VisualNow software.

Figure 3

MCC-IMS topographic plot of S. cerevisiae in early stationary phase during batch fermentation. Boxes indicate analytes with the most significant changes during the growth (perturbation) experiments.

Figure 4

(A) Fermentation profile of S. cerevisiae during batch growth in glucose minimal medium, (B) trends in intensity, (C) heat map of selected peaks detected by MCC-IMS analysis of the fermentation off-gas. Areas in the heat map show the detected analyte peak and the surrounding area; DO = dissolved oxygen.

Figure 5

(A) Fermentation profile of S. cerevisiae adh1Δ during batch growth in glucose minimal medium and (B) trends in intensity and (C) heat map of selected peaks detected by MCC-IMS analysis of the fermentation off-gas. The areas in the heat map show the detected analyte peak and the surrounding area; DO = dissolved oxygen.

Figure 6

MCC-IMS topographic plot of the off-gas of a glucose-limited continuous cultivation of S. cerevisiae. Boxes indicate analytes with the most significant changes during the growth (perturbation) experiments. The reaction ion peak (1/K₀ = 0.5 Vs cm⁻²) was compensated by the VisualNow software.

Figure 7

(A) Fermentation profile of S. cerevisiae during growth in a glucose-limited chemostat and (B) trends in intensity and (C) heat map of selected peaks detected by MCC-IMS measurements of the fermentation off-gas after perturbation of the metabolic steady state with a pulse of 22 mmol glucose; DO = dissolved oxygen.

Figure 8

(A) Fermentation profile of S. cerevisiae during growth in a glucose-limited chemostat and (B) trends in intensity and (C) heat map of selected peaks detected by MCC-IMS measurements of the fermentation off-gas during transition to anaerobic conditions; DO = dissolved oxygen.