Technical Abstract: N-caffeoyltryptamine is a compound belonging to a group of phenylpropanoid amides found in plants. For this study, N-caffeoyltryptamine was chemically synthesized and a high performance liquid chromatography (HPLC) method was developed for quantifying N-caffeoyltryptamine in biological samples. The synthesis was simple, and the yield of N-caffeoyltryptamine was greater than 50%. Using synthesized N-caffeoyltryptamine as a standard, HPLC separation was performed on a Nova-Pak C18 column using an isocratic buffer, and separation was performed using a coulometric electrochemical detector with four electrode channels. The detection of N-caffeoyltryptamine yielded an excellent peak resolution at a retention time of 18 min. The lower limit of the detection was as little as 100 fmol. Using this HPLC method, the plasma concentrations of N-caffeoyltryptamine were determined in mouse blood, collected at 5, 10, 15, 20, 25, 30, and 35 min following oral administration (1 and 3 mg/30 g body weight). This HPLC method, standardized with N-caffeoyltryptamine, is the first reported method able to quantify N-caffeoyltryptamine in standard and plasma samples with a good detection limit and consistent reproducibility.