Determination of carbohydrates present in Saccharomyces cerevisiae using mid-infrared spectroscopy and partial least squares regression

Abstract

A fast and simple method to control variations in carbohydrate composition of Saccharomyces cerevisiae, baker's yeast, during fermentation was developed using mid-infrared (mid-IR) spectroscopy. The method allows for precise and accurate determinations with minimal or no sample preparation and reagent consumption based on mid-IR spectra and partial least squares (PLS) regression. The PLS models were developed employing the results from reference analysis of the yeast cells. The reference analyses quantify the amount of trehalose, glucose, glycogen, and mannan in S. cerevisiae. The selection and optimization of pretreatment steps of samples such as the disruption of the yeast cells and the hydrolysis of mannan and glycogen to obtain monosaccharides were carried out. Trehalose, glucose, and mannose were determined using high-performance liquid chromatography coupled with a refractive index detector and total carbohydrates were measured using the phenol-sulfuric method. Linear concentration range, accuracy, precision, LOD and LOQ were examined to check the reliability of the chromatographic method for each analyte.

Figure

Comparison of workflows for carbohydrate determination in S.cerevisiae by FT-IR spectroscopy and HPLC-RI

Fig. 1

Diagram of the sampling and pretreatment steps of real samples (a); experimental process to carry out the reference analyses (b)

Fig. 2

Chromatograms of mannose using a mannan standard solution (a); trehalose and glucose (b) and glucose from glycogen (c) using a yeast standard sample

Fig. 3

FT-IR spectra of whole S. cerevisiae. Samples were drawn from nitrogen limit fed-batch fermentation 1 at different times over the course of the fermentation

Fig. 4

Variations of normalized total carbohydrate content for real samples from nitrogen limited fed-batch fermentation 1 of S. cerevisiae applying reference and FT-IR methods

Fig. 5

Relative trehalose, mannan, and glycogen content of the samples drawn from fermentations 1 and 2 at different times determined by reference analysis (black squares) and from FT-IR spectra by cross-validated PLS regression (red circles)