The serum concentrations, urinary and biliary excretions of six penicillin derivatives including aspoxicillin (ASPC) were studied in rats and the correlation between the values of pharmacokinetic parameters thus obtained and the Rm values measured by means of reversed phase TLC were analyzed. Among the penicillins studied, the hydrophilicity of amoxicillin was the highest (the lowest Rm value), which was followed by those of ASPC, ampicillin, p-hydroxypiperacillin, dehydroxyaspoxicillin and piperacillin in descending order. These Rm values were then correlated with the AUC values at 20 mg/kg of dosing, giving the results that more hydrophilic penicillins having a hydroxyl group show higher serum concentrations as well as greater AUC values. The studies of correlation between the Rm values and the urinary or biliary excretion revealed that hydrophilic penicillins were almost excreted into urine, but more hydrophobic ones were mainly eliminated into bile. From the above results, a hydroxyl group introduced to the phenyl group of ASPC was considered to have a role that increases the hydrophilicity of ASPC, giving higher and longer persistency of the serum levels and increasing the excretion of drugs into urine.