Simultaneous determination of free and N-acetylated polyamines in urine by semimicro high-performance liquid chromatography using 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride as a fluorescent labeling reagent.
We have developed a simple and highly sensitive semimicro high-performance liquid chromatographic method for the simultaneous determination of free and N-acetylated polyamines in urine. Polyamines and N-acetylated polyamines were derivatized with 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride to produce fluorescent sulfonamides. The labeling reaction was carried out at 50 degrees C for 15 min at pH 9. The fluorescent derivatives were separated on a reversed-phase column with a gradient elution using water-acetonitrile-methanol at 50 degrees C and detected by fluorescence measurement at 318 nm (excitation) and 406 nm (emission). The detection limits (signal-to-noise ratio=3) of the polyamines and N-acetylated polyamines were 0.7-4.5 fmol/injection. The within-day and day-to-day relative standard deviations were 3.2-7.9 and 3.0-7.7%, respectively. Significant differences were found in the urinary excretion of polyamines between cancer patients and normal subjects.
Inoue
Hirofumi
H
Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Fukuyama, Hiroshima 729-0292, Japan.
Fukunaga
Keiko
K
Munemura
Sayaka
S
Tsuruta
Yasuto
Y
D000107:Acetylation; D000328:Adult; D000368:Aged; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005260:Female; D005456:Fluorescent Dyes / Q000032:analysis*; D006801:Humans; D008297:Male; D008832:Microchemistry / Q000379:methods; D008875:Middle Aged; D009369:Neoplasms / Q000652:urine; D010797:Phthalimides; D011073:Polyamines / Q000652:urine*; D012016:Reference Values; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
C117624:4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride; D005456:Fluorescent Dyes; D010797:Phthalimides; D011073:Polyamines