Enzymatic Analysis of Yeast Biomass

Freeze-dried yeast biomass from experimental cultures was analyzed for the content of total glucans, β-(1,3)/(1,6)-glucan and α-glucan using the Mushroom and Yeast beta-glucan Assay Procedure K-YBGL enzymatic test (Megazyme). Based on the glucan content in biomass and the biomass yield, the productivity of β-(1,3)/(1,6)-glucan in the experimental cultures was determined and expressed in [g/L]. The content of total sugars was analyzed using the DNS method after prior acid hydrolysis of the biomass using 72% sulfuric acid. Nitrogen was determined using the Kjeldahl method (approx. 5-h mineralization of 50 mg of sample in the KjelFlex K-360 (Buchi) with the presence of 96% sulfuric acid and the Kjeltabs CT/3.5 catalyst. After distilling off the nitrogen, automatic titration of the samples was carried out using the TitroLine 5000 device (SI Analytics) and standard 0.1 M HCl solution. Nitrogen was converted to protein using the factor 6.25. The trehalose content was determined using the Trehalose Assay Procedure K-TREH enzyme test, Megazyme. The biomass (20 mg) was lysed in the presence of 0.5 mL of Y-PER reagent (Thermo Scientific) to release trehalose. The samples were incubated for 1 h at room temperature, while vortexing the samples several times. After incubation, the samples were centrifuged for 3 min at 15,000 RPM (MiniSpin Plus EPPENDORF). The supernatant was then collected in an appropriate amount to perform trehalose determinations in accordance with the test manufacturer's instructions.