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The biologically active forms of the isoflavones, aglycones (IA), in soy are reported to have many health benefits and could be considered as a “natural component” to replenish the estrogens in woman at menopausal and post-menopausal age. However, the isoflavones in soy exist principally in isoflavone glycoside (IG) forms, which have lower bioavailability. In order to improve health status of soymilk, it is essential to transform IG to IA. Initially, pure β-galactosidase and β-glucosidase were utilised for hydrolysing IG to IA in soymilk (SM). The level of hydrolysis ranged from 43.3-77.2% and 86.7-93.0% by various β-galactosidase concentrations and β-glucosidase, respectively. Six strains of probiotic organisms that produced β-galactosidase and β-glucosidase were used for the biotransformation of IG to IA in soymilk. To enhance the biotransformation level of IG to IA, lactulose and skim milk powder (SMP) were added to SM. The presence of lactulose in the medium enhanced the biotransformation level of IG to IA by Lactobacillus up to 21.9%. In particular, L. acidophilus 4461 biotransformed 88.8% IG to IA, the highest level recorded, in SM supplemented with lactulose 0.05% (w/w) (SML). The biotransformation of IG to IA was also enhanced significantly by 6.8 – 17.1% and 12.8 – 13.5% in SML by B. animalis subsp. lactis bb12 and B. longum 20099, respectively. Similarly, the biotransformation level of IG to IA in SM supplemented with SMP (SSM) ranged from 81.4 to 85.1%, which was 13.9 to 19.0% higher than that for SM. The levels of biotransformation were 84.0% and 85.4% for Bifidobacterium animalis subsp. lactis bb12 and B. longum 20099, respectively, compared to 74.3% and 72.8% for the SM. The supplementation with lactulose or SMP also significantly (P