The analytical data for identifying an unknown substance that was found in a nutrition supplement is presented. The unknown substance is purified using thin-layer chromatography and then measured using high-resolution mass spectrometry (HRMS) giving the exact mass from which the structure of the unknown substance was proposed. The procedure for synthesizing N-nitrosofenfluramine from fenfluramine is described. The extracted, synthesized, and standard N-nitrosofenfluramine are compared using HRMS, high-performance liquid chromatography (HPLC)-MS, HPLC-UV, Fourier transform IR spectroscopy, gas chromatography-MS, TLC, and NMR (1H NMR and 13C NMR). All analytical data obtained confirm that the unknown peak in the nutrition supplement is N-nitrosofenfluramine and that the synthetic procedure described can easily provide the N-nitrosofenfluramine reference substance for identification.