Theoretical considerations have suggested that variations in the resistance of the unstirred water layer (UWL) have a profound effect on the kinetic constants of intestinal transport. In this study, a previously validated in vitro technique was employed to determine the unidirectional flux rate of glucose, galactose, 3-O-methyl glucose and fructose into the rabbit jejunum under carefully-defined conditions of stirring of the bulk phase known to yield different values for the effective resistance of the UWL. For each monosaccharide, uptake is much greater when the resistance of the UWL is low than when high. The maximal transport rate, Jd m, of glucose was half as large as the Jd m of galactose and 3-O-methyl glucose (3-O-MG), and was twice as great as the Jd m of fructose. The apparent affinity constant, Km * of glucose is less than that of fructose, which was lower than the Km * of galactose and 3-O-MG. The use of the Lineweaver-Burk double reciprocal plot is associated with an overestimation of both Jd m and Km *. This discrepancy between the true and apparent values of the kinetic constants is much greater for lower than for higher values of Jd m and Km *; variations in the resistance of the unstirred layer influences the magnitude and direction of the discrepancy. The apparent passive permeability coefficient is similar for each sugar, but because of the different values of Jd m, passive permeation contributes relatively more to the uptake of glucose and fructose than of galactose or 3-O-MG. Under conditions of high unstirred layer resistance, differences in uptake rates of the sugars are due to differences in their Jd m rather than their Km *. Kinetic analysis is compatible with the suggestion that the glucose carriers are predominantly near the tip of the villus, whereas those for galactose and 3-O-MG are located along the entire villus and the Km * of their carriers at the tip is lower than their Km * towards the base of the villus. It is proposed that there are multiple or heterogeneous intestinal carriers for glucose, galactose and 3-O-methyl glucose in the jejunum of the rabbit.