Arylamine N-acetyltransferase (NAT) activity in a human liver tumor (heptoma) cell line was inhibited by vitamin C. Using high performance liquid chromatography, NAT activity on the acetylation of 2-aminofluorene and p-aminobenzoic acid was examined. Two assay systems were performed, one with cellular cytosols, the other with intact liver tumor cell suspensions. The NAT activity in a human liver tumor cell line was inhibited by vitamin C in a dose-dependent manner in both types of examined system: i.e. the greater the concentration of vitamin C in the reaction, the greater the inhibition of NAT activities in both systems examined. The data also indicated that vitamin C decreased the apparent Km and Vmax of NAT enzymes from human liver tumor cells in both systems examined. This report is the first demonstration which showed vitamin C effect on human liver tumor cell NAT activity.