Determination of concentration of living immobilized yeast cells by fluorescence spectroscopy

Fluorescence spectroscopy is one of suitable methods for the direct monitoring of immobilized cells because it does not require (in certain arrangement) transparent environment like absorbance measurements. The aim of this work was to develop a method allowing measurements of concentrations of living immobilized cells using only intrinsic fluorescence of biogenic fluorophores in cells. Saccharomyces cerevisiae yeast cells were immobilized into alginate and into a mix of alginate with prepolymerized tetramethoxysilane (TMOS) and measured in flow through cuvettes. Transitions from aerobic to anaerobic conditions were made by stopping circulation of perfusing medium and 2-dimensional fluorescence (2-DF) spectra and time responses of nicotinamide adenine dinucleotide coenzymes (NAD(P)H) fluorescence intensity were measured. Fluorescence intensity of intracellular NAD(P)H, its changes during aerobic-anaerobic transitions and the time delay between halt of medium circulation and rise of NAD(P)H fluorescence intensity correlated with concentration of living immobilized yeast cells. In case of alginate–TMOS layers, only relative change of NAD(P)H fluorescence intensity corresponded to the content of active biomass in the whole range of measured concentrations.