抄録
A simple, rapid and colorimetric method for the estimation of histamine, based on the extraction of histamine azo-compound with ethyl acetate, was presented. Coloration of histamine with diazo-reagent in ice bath, was stable and reproducible, and the intensity of histamine was proportional to the concentration of histamine, showing linear relation within the range from 1 to 20γ. Many well-known substances, in the amount that might be contained in fish muscle, scarcely interfered with the method. Histamine added to fish extract was recovered with satisfactory results. Applications on several species of fish muscle showed as good results as those obtained by other recent method.
Analytical procedure was summarized as follows:
Fish muscle extract was obtained by shaking and filtering, after being added succesively 10 volumes of water and of 5% CCl3COOH. One cc. of the extract was neutralized with 0.5% NaOH and made to 2 cc. with water and then 1 cc. of 4% Na2CO3 was added to it. To the above solution, placed in ice bath over for 3 minutes, was added 1 cc. of ice-cooled diazo-reagent and the solution was kept in ice bath. After cooling it for 3 minutes almost exactly, 7 cc. of ethyl acetate was added to it and its solution was shaken vigorously for approximately 30 seconds. After brief standing, almost all of ester layer was taken and shaken with 5 cc. of about 0.1% Na2CO3. After letting it stand, upper portion was taken and dried with granular anhydrous Na2SO4. Rose color of ester solution was immediately measured in colorimeter (filter 500mμ, cell 10mm.). Diazo-reagent was freshly prepared by adding 0.2 cc. of 5% NaNO2 to 10 cc. of ice cooled 0.1% p-nitroaniline in 0.1 N HCl solution and in advance of its use it had to be kept in ice water.
