Pre-column Derivatization-HPLC-MS Determination of Free Polyamines in Urine with Fluorescence Detection

A new fluorescence reagent,1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate was used as pre-column derivatizing agent in an acetonitrile solution,catalyzed by the borate buffer of pH 9 to react with polyamines at 40 ℃ for 10 min,to give stable fluorescence products.Five polyamines were then separated on the Eclipse XDB-C8 chromatographic column by gradient elution with acetonitrile-H2O solution of different concentrations,and on-line analyzed qualitatively with APCI mode by MS and quantitatively by HPLC with fluorescence detection at the wavelengths of (λex) 333 nm and (λem) 390 nm.Detection limits of the 5 polyamines (3S/N) found were in the range of 4.98-9.31 fmol.Test for recovery was made by addition of standard polyamine solutions to a urine sample,and the values of recovery found were in the range of 94%-98%.