Glycemic Index (GI) measures how carbohydrate-containing or sugary food increases blood glucose. Consumption of high GI food has been associated with many chronic conditions like obesity and diabetes; hence, monitoring of dietary GI is important. However, the standard method to determine GI is tedious and costly as it uses human subjects. This study investigated the possible utilization of yeast (Saccharomyces cerevisiae) fermentation system in ex vivo determination of sugar GI. Clonal S. cerevisiae were incubated in separate media containing known concentrations of test brown and white sugars and glucose as reference sugar. The concentrations of the unfermented sugars were measured at 25-minute interval for 100 minutes to plot the yeast response curve to each sugar. Using the standard incremental area under curve (IAUC) and GI formula, the calculated GI for brown, 125, was lower than the calculated GI for white sugar, 132, using the yeast system. This sequence is consistent with the sugars’ published GIs obtained using human subjects. Hence, S. cerevisiae may be developed as a simple alternative system in measuring food GI. Future biotechnology work on S. cerevisiae carbohydrate metabolism and mathematical remodeling of the IAUC and GI formula to suit GI determination using S. cerevisiae system are recommended.