A modified high-performance liquid chromatographic method for the quantitative determination of 6-methoxy-2-naphthylacetic acid (6-MNA), a major metabolite of nabumetone, in human serum was developed and validated. The composition of the mobile phase in Daigneault & Ferslew's method was changed in this study to improve the separation of 6-MNA from serum components. The volume ratio of acetonitrile to 1.5% acetic acid solution was changed from 60: 40 to 25: 75 in this study. As a result of the modification, the separation of 6-MNA from 2-naphtol (internal standard) and serum components was greatly improved. At a flow rate of 3.0 ml/min of the mobile phase, retention times of 6-MNA and 2-naphtol were 13 and 9 min, respectively. The detection limit was 1 μg/ml. Intra- and interday variations of the assay were <10.0 and <7.28%, respectively. Intra- and interday relative errors were <7.82 and <6.76%, respectively. 6-MNA in human serum could be determined successfully after oral administration of nabumetone (1g). Thus, the modified UV-HPLC method was confirmed to be applicable to the pharmacokinetic study of 6-MNA after oral administration of nabumetone to human subjects.