Development of Ultra-sensitive HPLC Determination Method for Histamine Based on Intramolecular Excimer-forming Fluorescence Derivatization and Its Applications to Clinical and Neuroscience Investigations

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Sensitive and selective HPLC determination methods for histamine related compounds based on intramolecular excimer-forming fluorescence derivatization have been developed. At first, the suitable reagent was screened including the design of novel reagents, and derivatization and HPLC conditions were optimized for the preliminary examinations using histamine standard. By the validation tests, the proposed method is more selective and sensitive than the previous methods with tedious pretreatment procedure. Therefore, the following application studies were carried out.(1) Human urine ; the amounts of histamine in only 1 μL of human urine were determined by the established method with high accuracy without any pretreatment. (2) Rat plasma and tissues; the histamine levels in rat plasma and tissues (abdominal skin, lung, stomach, heart and liver) were measured using excimer fluorescence derivatization method after a simple pretreatment. The obtained values in plasma and tissues were in good agreements with those reported by other workers. (3) Microdialysis samples from rat brain ; real-time monitoring of histamine in rat brain could be carried out by combined with microdialysis techniques. The relationships between the amounts of histamine in some regions of brain (hypothalamus, prefrontal cortex, hippocampus and striatum) and the influence of drugs were investigated.At last, simultaneous determination method of histamine and histidine (precursor of histamine) based on intramolecular excimer-forming fluorescence derivatization was developed. This method was highly sensitive and selective, and useful for clinical and food sanitary fields.In conclusion, the present derivatization technique allows simple, selective and sensitive determination of histamine related compounds.