The brain uptake index (BUI) of polar 14C-labeled test compounds with molecular weights (MWs) of 79-70,000 was examined using the single-pass carotid injection technique in pentobarbital-anesthetized rats. Compounds were injected in 40 mM malonate, pH 2.5, and 10 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid, pH 7.4. BUI is defined as (14C-labeled test compound/[3H]H2O in brain)/(14C-labeled test compound/[3H]H2O in injectate) x 100 at a 5-s decapitation time. Increased BUIs of compounds in pH 2.5 buffer were MW dependent with a threshold < 5,000. BUI, extraction, and permeability-surface area product (PS) were significantly greater at pH 2.5 compared with pH 7.4 (P < 0.05). Washouts of [14C]butanol and [3H]H2O at pH 2.5 and pH 7.4 were calculated. Cerebral blood flow and PS increased at low pH. Other buffers, oxalate, glycine, and lactate were used at low pH and also increased BUIs. The duration of the blood-brain barrier (BBB) opening at pH 2.5 was 60 s as estimated by penetrance of the normally excluded dye fluorescein. A plot of BUI or PS at pH 2.5 vs. (MW)1/2 suggests that transient BBB tight junction opening contributed to the passage of polar compounds at low pH.