High-performance liquid chromatography of plasma catecholamines using 1,2-diphenylethylenediamine as precolumn fluorescence derivatization reagent.
A simple, rapid and highly sensitive method for the determination of catecholamines (norepinephrine, epinephrine and dopamine) in human plasma is described which employs high-performance liquid chromatography with fluorescence detection. After cation-exchange chromatography on a Toyopak SP cartridge, the catecholamines and isoproterenol (internal standard) in 500 microliters of plasma are converted into the corresponding fluorescent compounds by reaction with 1,2-diphenylethylenediamine in aqueous acetonitrile. These compounds are separated within 8 min on a reversed-phase column, TSK-gel ODS-120T, with isocratic elution using a mixture of water, methanol and acetonitrile containing a Tris--hydrochloric acid buffer (pH 7.0). The detection limit for each catecholamine is ca. 2 fmol in a 100-microliters injection volume. N-Methyldopamine can also be used as internal standard.
D000328:Adult; D000537:Aluminum Oxide; D002395:Catecholamines / Q000097:blood*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D005029:Ethylenediamines; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D007545:Isoproterenol / Q000032:analysis; D008297:Male; D008875:Middle Aged; D010472:Perchlorates; D013050:Spectrometry, Fluorescence; D014462:Ultrafiltration
D002395:Catecholamines; D005029:Ethylenediamines; D007202:Indicators and Reagents; D010472:Perchlorates; D007545:Isoproterenol; D000537:Aluminum Oxide; C007305:1,2-dianilinoethane