This study examined the effects of plasma α1‐acid glycoprotein (AGP) on the protein binding and pharmacokinetics of trimethoprim (TMP) in piglets. The piglets were given 5 mg/kg of TMP intravenously at 1, 14 and 28 days after birth. The plasma AGP level was highest at day 1. Fourteen days after birth, the level decreased by about 90% of that at day 1. The level at 28 days was almost the same as that at 14 days. Plasma protein bindings of TMP depended on the AGP level but not on the albumin level. The percentage of plasma protein binding decreased from 85 to 45%, and the AGP level also decreased from 6,000 to 700 μg/ml. The altered protein binding of TMP affected pharmacokinetic parameters such as total body clearance (CLtot), distribution volume and therefore the elimination rate constant. These parameters correlated well with the percentage binding to plasma proteins. Maturational development in the capacity to eliminate TMP was also indicated by the increase in total body clearance of unbound drug (CLtotub), which directly reflects the elimination capacity of the body. However, its contribution to the increase in CLtotwas considered not to be large. CLtotubincreased twofold 14 days after birth, whereas CLtotincreased about ninefold. The increase in CLtottherefore, may result from both the maturational development in elimination capacity and the AGP‐dependent decrease in plasma protein binding. It is concluded that the decrease in plasma AGP level obsered in piglets is one of major factors affecting the pharmacokinetics of TMP.