Simultaneous determination of histamine and N alpha-methylhistamine in biological samples by an improved enzymatic single isotope assay.

Abstract

A modified highly sensitive and specific radioenzymatic assay for the simultaneous determination of histamine (HA) and N alpha-methylhistamine (N alpha-MH) in tissues and body fluids is described. In the presence of the enzyme histamine-N-methyltransferase and of the methyl donor [3H]-S-adenosylmethionine, the transmethylation process was about seven times more effective for HA than for N alpha-MH. In the same conditions only very low amounts of N alpha, N alpha-dimethylhistamine (N alpha, N alpha-DMH) were converted into its 1-methylated derivative. The high degree of specificity attained by this method is due to the rapid quantitative extraction of the biological fluids, to the partially purified enzyme preparation and to the thin-layer chromatography system used which allows an excellent separation of the 3H-1-methyl products of HA, N alpha-MH and N alpha, N alpha-DMH. This method is highly sensitive for the assay of HA and N alpha-MH (detection limit 10 and 50 picograms, respectively), but due to lack in sensitivity, it cannot be extended to the measurement of N alpha, N alpha-DMH. The HA content of 20-30 samples can be determined in duplicate by one person in a working day. The concentrations of HA measured by this method in different biological samples (human whole blood, plasma, urine, gastric juice and skin biopsies) were in good agreement with the values reported in the literature. The presence of minute amounts of N alpha-MH in the human gastric juice was established by rigorous checking.