An attempt had been made to isolate and characterize yeast strain from different saccharine materials viz, apple juice, pineapple juice, mango juice, mosambi juice, orange juice, pomegranate juice and Lichi juice. A total of 7 samples from different kind of fruit juices sources were isolated using selective medium i.e., Yeast Extract Peptone Dextrose Agar (YEPDA). Differential tests were applied including morphological, cultural and biochemical characteristic, which facilitate the opportunity for identification of the yeasts. The strains were tested for growth at different temperature to check their mortality. These strains were further screened and tested on molasses for osmo-tolerant properties. Activated Dry yeast was taken as control. The performance of isolated yeast strains was tested on molasses. Out of different fruit juices strains obtained from apple and orange gave the best results in term of ethanol content 10.28 % & 10.15% and also Fermentation Efficiency 89.39 % and 88.26% respectively which was at par with ADY. Moreover, the yeast strains were also tested for its osmo-tolerance and thermo-tolerance using molasses as raw material for production of ethanol. Result revealed that total reducing sugar (TRS) of 14%, 18%, 21% at all these concentrations the isolated strain growth was observed however apple and orange isolates gave best result in terms of ethanol yield