Significant amounts of entrapped oil could be potentially recovered from water-flooded reservoirs nearing their economic limit of production via biosurfactant-enhanced oil recovery. However, evidence for the persistence and metabolic activity of injected biosurfactant-producing bacterial species activity is lacking. We injected a glucose-nitrate-mineral nutrient mixture and two lipopeptide biosurfactant-producing Bacillus strains into two wells to correlate in-situ metabolism with oil recovery. Two wells producing from the same Viola formation were each inoculated with about 60 m 3 of tank battery brine with a nutrient mixture containing glucose, sodium nitrate and trace metals and Bacillus licheniformis RS-1 and Bacillus subtilis subsp. subtilis spizizenii NRRL B-23049. Analysis of production water indicated in-situ growth of the injected strains and other heterotrophic fermenting bacteria, metabolism of the nutrients, and biosurfactant production. Both wells had a peak lipopeptide biosurfactant concentration of 20 and 28 mg/L, respectively, which is twice the minimum concentration required to mobilize entrapped oil from sandstone cores. Metabolic products of glucose fermentation in both wells were acetate, 2,3-butanediol, ethanol, formate, lactate, and succinate and cells and these products accounted for 107.6% of the glucose used. The increase in biosurfactant, acids, alcohols and carbon dioxide during the first 5 days after commencement of production corresponded directly with an increase in oil recovery. Wellhead measurements of total produced fluid, the water-to-oil ratio and the percentage of oil showed that about 52.5 m 3 of additional oil (net cumulative increase) occurred during the first 60 days of sampling. These results showed the feasibility of stimulating in-situ biosurfactant production and its potential to improved oil production from mature oil reservoirs.