HPLC determination of polyamines in urine

Determination of polyamines (putrescine, spermidine and spermine) in urine samples of patients was worked out using high performance liquid chromatography (HPLC) and pre-column derivatization. For HPLC separation, a method of reversed-phase chromatography with gradient elution of water-organic mobile phases, UV and fluorescence detection were applied. Special care was devoted to the preseparation step, the hydrolysis, derivatization and selective extraction of clinical samples. The optimal HPLC conditions were recommended for simultaneous determination of putrescine, spermidine and spermine in a minimal analysis time (about 15 min) and suitable chromatographic resolution values (Rij greater than 1.5). Detection limits for all polyamines were evaluated (40 pmol for putrescine, 25 pmol for spermidine and 20 pmol for spermine) for the analysis of polyamines in the urine. The urine samples of healthy people and the cancer patients were compared and the procedure or the complete analysis has been described.