Determination of Viable Yeast Cells by Gravitational Field‐Flow Fractionation with Fluorescence Detection

Abstract

Vinification processing is largely related to yeast performance and depends on the initial cell viability. To optimize the quality of wine fermentation, control of the yeast quality is mandatory. The present paper describes a new method using gravitational field flow fractionation (GrFFF) with fluorescence detection for the determination of yeast cell viability before the fermentation process. A GrFFF calibration procedure was developed using commercial yeast to prepare standards of viable cells and propidium iodide (PI) as fluorescent probe for nonviable cells. The suitability of the new method was tested with several commercial yeast strains with a g/L content ranging from 1 to 3. The validation of the method was performed by comparing GrFFF viability values with those obtained using Coulter counter and flow cytometry techniques.

Kinetic Study of Cell Proliferation of Saccharomyces cerevisiae Strains by Sedimentation/Steric Field Flow Fractionation in Situ

“…Comparing the size distributions of the yeast cells obtained by the above two techniques in diploid cells (or clusters and chains), the differences that appeared can be justified by the abnormal shape of the analyte particles. Nevertheless, the final results for the "large" cells, although different from those obtained by SEM, are in good agreement with the literature (5,9), always in respect to the standard deviations.…”

“…According to the results, the S. cerevisiae strains show a bimodal size distribution with weight average diameters in the range of 3.5-8.6 µm, which are in good agreement with those sizes mentioned in the literature (total cells counted from 2-7 µm) (5,9). Specifically, Sd/ StFFF is an analytical technique that can be used with great accuracy for the determination of the single yeast cells particle size distribution, as it occurs from the comparison studies with SEM.…”

“…The forms of these fractograms indicate a bimodal size distribution of S. cerevisiae strains immediately after the disperse of the cells into the culture medium, which is in full agreement with Coulter counter data obtained by Sanz et al (5,9). By the time the yeast cells begin to consume the culture medium, their metabolic activities are in full development.…”

“…Classical biochemical methods are not suitable for characterization and kinetic study of the S. cerevisiae strains (2). In the past few years the Field Flow Fractionation technique has been applied for the characterization of yeast cells (3)(4)(5)(6)(7), as well as the study of the viability and activity of S. cerevisiae strains during wine fermentation (8,9).…”

Use of Fluorescent Probes for Determination of Yeast Cell Viability by Gravitational Field-Flow Fractionation

“…Among the techniques in this family, gravitational field-flow fractionation (GrFFF) is the experimentally simplest FFF mode and an inexpensive tool for the successful separation, characterization, and micropreparation of different particles (16)(17)(18)(19). As a result of its mild experimental requirements, GrFFF provides unique separation capabilities for biological samples and has recently been applied for the characterization of several yeasts samples (20)(21)(22)(23)(24)(25)(26)(27)(28). The FFF technique is a very versatile one because it can be coupled with different types of detection systems, such as UV, light scattering, flow cytometry (27), or chemiluminiscence (23,24).…”