We tested two hypotheses to account for the reduction in coupling of anionic solute to water flow (solvent drag) in microvessels during perfusion with plasma compared with albumin. Solvent drag is determined by both hydraulic conductivity (Lp) and solute reflection coefficient (sigma). Accordingly, decreased solvent drag during plasma perfusion must be the result of an increase in sigma (hypothesis 1) or reduction of Lp (hypothesis 2) or some combination of both mechanisms. These hypotheses were assessed by measuring Lp, sigma, and diffusive solute permeability (Psd) to the anionic protein alpha-lactalbumin in frog mesenteric exchange microvessels during plasma or albumin perfusion. The solute permeability coefficient to alpha-lactalbumin (Ps alpha-lactalbumin) was lower during exposure to plasma than bovine serum albumin (BSA) [(Ps alpha-lactalbumin)plasma/(Ps alpha-lactalbumin)BSA = 0.31 +/- 0.11 (means +/- SE, n = 9)]. Solute reflection coefficient to alpha-lactalbumin (sigma alpha-lactalbumin) was 0.69 +/- 0.02 in plasma and 0.34 +/- 0.03 in BSA (n = 7). Lp was not significantly influenced by perfusate protein composition (Lp plasma/Lp BSA = 1.02 +/- 0.11; n = 20). These data lead to the conclusion that the actions of plasma are to confer charge selectivity for anionic solute and, to a lesser extent, modify the porous pathways of the microvessel wall. Taken together, these results indicate that porous pathways contribute significantly to macromolecular flux in plasma-perfused vessels.