Simultaneous determination of 3,4-dihydroxyphenylacetic acid and homovanillic acid using high performance liquid chromatography-fluorescence detection and application to rat kidney microdialysate.
We established a sensitive and simultaneous determination method of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) using HPLC-fluorescence detection. This method adopted the column-switching system, which included an on-line extraction of carboxylic acids by a strong anion-exchange column followed by separation on an ODS column, coulometric oxidation, fluorogenic reaction with ethylenediamine, and fluorescence detection. The detection limits were 50 and 100 fmol/injection for DOPAC and HVA, respectively (a signal-to-noise ratio of 3). The method was applicable to 50 microl of rat kidney microdialysate with a sufficient accuracy and precision. The concentrations of DOPAC and HVA in rat kidney microdialysate were 131+/-29 and 404+/-44 nM, respectively (n=5). This is the first report of DOPAC and HVA quantified in rat kidney microdialysate.
Laboratory of Bio-Analytical Chemistry, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo, Japan.
Makoto
M
Tsunoda
Kazuto
K
Mitsuhashi
Mayumi
M
Masuda
Kazuhiro
K
Imai
D015102:3,4-Dihydroxyphenylacetic Acid / Q000032:analysis*; D000818:Animals; D000837:Anion Exchange Resins / Q000737:chemistry; D002264:Carboxylic Acids / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D005029:Ethylenediamines / Q000737:chemistry; D005453:Fluorescence; D006719:Homovanillic Acid / Q000032:analysis*; D007668:Kidney / Q000737:chemistry*; D008297:Male; D017551:Microdialysis; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D014020:Tissue Extracts / Q000737:chemistry
D000837:Anion Exchange Resins; D002264:Carboxylic Acids; D005029:Ethylenediamines; D014020:Tissue Extracts; D015102:3,4-Dihydroxyphenylacetic Acid; C031234:ethylenediamine; D006719:Homovanillic Acid