Single pump column switching technique employing a flow gradient and wavelength programmed fluorescence for simultaneous monitoring of serotonin, fluoxetine and norfluoxetine in rat brain microdialysate.
A single pump column switching technique with multidimensional chromatography, flow gradient and wavelength programmed fluorescence detection was developed for simultaneous quantitation of serotonin, fluoxetine and norfluoxetine in rat brain microdialysate. The column switching was configured such that position I of the switching valve employed column I (50 mm length) and column II (250 mm length) in series. This configuration resulted in optimal resolution of serotonin from interfering neurochemicals from rat brain. After elution of serotonin at 13.2 min the valve was switched to position II in which the flow of the mobile phase was directed through column I only. Flow gradient programming was then used to ramp the flow rate from 0.1 to 0.4 ml min-1 which resulted in optimal elution of fluoxetine and norfluoxetine. Strategic optimization of the single mobile phase enabled use of a single pump and detector making the analytical system simple and cost effective. Wavelength programmed fluorescence enabled sensitive detection of the analytes despite the difference in their fluorescence spectrum. The limit of detection for serotonin, norfluoxetine and fluoxetine were 10, 612 and 523 fmol, respectively. Rat brain microdialysate samples demonstrated selectivity for serotonin, fluoxetine and norfluoxetine. The method demonstrates application to the study of site specific neuropharmacokinetics and neuropharmacodynamics of fluoxetine in vivo.
Ramaiya
A
A
Department of Pharmacy and Pharmaceutics, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0533, USA.
March
C
C
Karnes
H T
HT
D000818:Animals; D002138:Calibration; D016903:Drug Monitoring / Q000379:methods*; D005473:Fluoxetine / Q000031:analogs & derivatives* / Q000032:analysis*; D008297:Male; D017551:Microdialysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012701:Serotonin / Q000032:analysis*
D005473:Fluoxetine; D012701:Serotonin; C036139:norfluoxetine