Simultaneous determination of histamine and N tau-methylhistamine with high-performance liquid chromatography using electrochemical detection.
We have developed a liquid chromatographic method which uses electrochemical detection for the simultaneous quantitation of histamine and N tau-methylhistamine in rat brain. The amines are derivatized with the water-soluble Bolton-Hunter reagent (sulfo B-H). Perchloric acid extracts of rat brains are chromatographed on a strong cation-exchange resin. The eluate is evaporated and allowed to react with sulfo B-H at pH 9.8 at room temperature. The derivatization is complete after 30 s vortexing. The derivatives are purified using a cellulose-phosphate fibrous cation exchanger. They are quantified with an electrochemical detector at a potential of 0.56 V after preoxidizing the sample at 0.47 V. The derivatives of histamine, N tau-methylhistamine, and N alpha-methylhistamine are completely separated without interfering peaks. Since no N alpha-methylhistamine was detected in rat brain it was used as an internal standard. The detection limits are 0.1 pmol of histamine and 0.2 pmol of N tau-methylhistamine. The precision of this method is high, with within-run and between-run coefficients of variation of 2-7% and linearity of 0.999. Both histamine and N tau-methylhistamine peak heights increased significantly and selectively after treatment with pargyline. Because of the high sensitivity, accuracy, and precision, the histamine and N tau-methylhistamine contents of single nuclei of the rat hypothalamus can be routinely quantified.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004563:Electrochemistry; D006632:Histamine / Q000032:analysis*; D007031:Hypothalamus / Q000032:analysis; D007093:Imidazoles; D008297:Male; D008761:Methylhistamines / Q000032:analysis*; D010293:Pargyline / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D013388:Succinimides
D007093:Imidazoles; D008761:Methylhistamines; D013388:Succinimides; C031205:3-methylhistamine; C022581:Bolton-Hunter reagent; D006632:Histamine; D010293:Pargyline; C048140:N-methylhistamine