Determination of Histamine by High-Performance Liquid Chromatography After Precolumn Derivatization with o-Phthalaldehyde-Sulfite

A fast and sensitive method was developed for in vivo determination of histamine in the brain microdialysate by reverse ion pair chromatography with electrochemical detection. The microdialysates were derivatized with o-phthalaldehyde and sodium sulfite, and separation was achieved using isocratic elution within 10 min. The separation was performed in an Agilent Eclipse Plus C18 column (3.0 x 150 mm, particle size 3.5 mu m), and the mobile phase consisted of 100 mM monosodium phosphate (pH 6.0), 500 mg L-1 OSA and 20% methanol (v/v). The linearity (R-2) was found to be >0.999, with a range from 2 to 50 nM and excellent repeatability (relative standard deviation, 2.29-6.04%), and the limit of detection was 0.4 nM. This method was successfully applied to analyze the extracellular concentration of histamine in the hypothalamus of rats, with probe recovery calculated in vivo.