In Vitro Glucose Uptake Assay in Yeast Cell

The commercial baker’s yeast, Saccharomyces cerevisiae was grown in YM medium (1% dextrose, 0.5% peptone, 0.3% malt extract, 0.3% yeast extract, 2% agar) and the culture was centrifuged at 3,000 × g for 5 min. The supernatant was discarded and yeast cell pellet was taken for the experiment. Further, a 10% (v/v) of yeast suspension was prepared in sterilized deionized water. Various concentrations of AEMOL (50–500 μg/mL) were added to 1.0 mL of glucose solution (5, 10, and 25 mM), separately and incubated together at 37°C for 10 min. Glucose uptake was initiated by adding 100 μL of yeast suspension in each extract-glucose solution. The mixture suspension was vortexed and further incubated at 37°C for 60 min. After 60 min of incubation, the suspension was centrifuged (3,000 × g, for 5 min) and the supernatant was used to estimate the amount of glucose present (Nair et al., 2013). In yeast kinetic assay, V_max and K_m were calculated for both controls as well as the treatment group. Detailed procedure for the yeast cell kinetic assay has been described in “kinetic study of ex vivo glucose uptake.” The percentage increase of glucose uptake in yeast cells by using AEMOL was calculated using the underneath formula:

Where, absorbance of control reaction containing all reagents excluding the test sample; absorption of sample reaction containing AEMOL.