Profiling of amine metabolites in human biofluids by micellar electrokinetic chromatography with laser-induced fluorescence detection.
A rapid and sensitive capillary electrophoresis (CE) method has been developed for profiling organic metabolites containing amine functional groups in mammalian biofluids. Metabolites containing an amine group were derivatized with 4-fluoro-7-nitrobenzo-2,1,3-oxadiazol (NBD-F), separated by micellar electrokinetic chromatography (MEKC), and detected by argon-ion (488 nm) laser-induced fluorescence detection. The optimized MEKC background electrolyte conditions were: 50 mmol L-1 sodium cholate, 5 mmol L-1 beta-cyclodextrin, and 20 mmol L-1 Brij 35 in 20 mmol L-1 aqueous borate buffer, pH 9.3, containing 7% methanol. Under these conditions all the amine compounds in mammalian biofluids, for example plasma, saliva, and urine, were derivatized directly, without extraction, in a minimum volume of 100 nL and the derivatives could be separated within 16 min. Up to 90% of the amine-containing metabolites in plasma and saliva could be identified by reference to standard compounds. For twenty amine standards linearity of calibration was better than R2=0.99. Migration-time and peak-area reproducibility were better than RSD 1.5% and 15% respectively. In replicate analysis of human plasma bioanalytical precision ranged between 0.7 and 3.8 RSD% for a 5.0-microL volume and between 1.7 and 5.5 RSD% for 100-nL volume. The concentrations measured were found to be in agreement with literature values.
Tseng
Hua-Ming
HM
Centre for Analytical Bioscience, School of Pharmacy, University of Nottingham, Nottingham, NG7 2RD, UK.
Li
Yin
Y
Barrett
David A
DA
D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives / Q000737:chemistry; D000588:Amines / Q000032:analysis* / Q000097:blood / Q000652:urine; D000596:Amino Acids / Q000097:blood / Q000737:chemistry; D001826:Body Fluids / Q000737:chemistry*; D002138:Calibration; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods; D006801:Humans; D007834:Lasers; D015203:Reproducibility of Results; D012463:Saliva / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000295:instrumentation / Q000379:methods*
D000588:Amines; D000596:Amino Acids; C032930:7-fluoro-4-nitrobenzo-2-oxa-1,3-diazole; D009327:4-Chloro-7-nitrobenzofurazan