Activation of intestinal Na⁺–glucose cotransport increases paracellular movement of inert tracers in cultured monolayers, isolated rodent intestinal mucosae, and in rodents in vivo. However, not all studies have demonstrated comparable effects on human intestinal paracellular absorption. We sought to assess the effects of Na⁺–glucose cotransport on paracellular absorption in human beings using a simple noninvasive assay. Study subjects drank six 200-ml doses of test solution, composed of 0.8% w/v creatinine (sufficient to overwhelm endogenous creatinine) in 277 mM glucose or mannitol and urine was collected. Intestinal creatinine absorption is paracellular. Once absorbed, creatinine is cleared into the urine. Therefore, urinary creatinine recovery reflects intestinal paracellular creatinine absorption. Total urinary creatinine recovery was 55% ± 4% of creatinine ingested with glucose and 38% ± 9% of creatinine ingested with mannitol (p < 0.001). Thus, intestinal paracellular absorption of creatinine is increased by the presence of luminal glucose. Our results are consistent with in vivo human regulation of mucosal permeability by Na⁺–glucose cotransport.