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Glyburide is a second-generation sulfonylurea hypoglycemic drug used for the treatment of select women with pregestational and gestational diabetes mellitus (GDM). In vitro and in vivo investigations demonstrated its very low transplacental transfer to the fetal circulation. However, the factors influencing its low transfer across the human placenta remain unclear. Therefore, the goal of the current investigation was to determine the effect of human serum albumin (HSA) on the transfer and distribution of glyburide across the human placenta. To achieve this goal, the technique of dual perfusion of the placental lobule was utilized. The effect of HSA on the transfer of glyburide was determined at the range of glyburide to HSA molar ratios of 1:2-1:100. The transfer rate of free/unbound glyburide to the fetal circuit was 73 ± 10% of the freely diffusible marker compound antipyrine (AP). Data obtained indicates the dependence of glyburide transfer and its retention by the placental tissue on the concentration of HSA.