A method is presented for determining the concentrations of several oxidized pterins in a variety of biological samples. This methodology employs reverse-phase "ion-pair" liquid chromatography with electrochemical detection. Detection limits below a picomole have been achieved with responses linear over four orders of magnitude. This methodology was employed to study the excretion pattern of pterins in human urine. The concentration of several pterins in mouse brain and liver samples were also determined. The direct detection of tetrahydrobiopterin was demonstrated.