A simple method for the determination of diamine oxidase (DAO) activity is described. The method is based on oxidation of histamine with DAO, formation of histamine-copper(II) chelated cation from the remaining histamine, and extraction of the ion associated with tetrabromophenolphthalein ethyl ester. A linear calibration curve between absorbance and DAO activity was obtained for DAO activity up to 250 mU/ml. The inter-assay coefficient of variation was 2.12% with a recovery of 101.7% at the DAO activity of 123.5 mU/ml. A close correlation was found between the results of this method and the 2,2'-azino-di(3-ethylbenzthiazoline-6-sulfonic acid) method.Keywords diamine oxidase; histaminase activity; colorimetry; solvent extraction; histamine-cupric ion-tetrabromophenolphthalein ethyl ester associate Diamine oxidase [diamine: oxygen oxidoreductase (diaminating), EC 1.4.3.6] (DAO) is widely distributed in animals and microorganisms." In animals, DAO is relatively abundant in the kidney,2) and is also present in the plasma and other cells. DAO activity has been considered a useful parameter in clinical studies.3) DAO activity has been determined by the use of radioactive substances,4,5) highperformance liquid chromatography6,7) (HPLC), and colorimetric methods.8-11) The first method is the most sensitive but required special equipment, and is not suitable for general use. The second method is time-consuming and troublesome. The third method is not as sensitive as the other methods, but is the simplest, and is suitable for routine use. In the colorimetric method, the reactions are based on DAO oxidase activity towards several short-chain diamines,11) such as ethylenediamine, putrescine (Pu), cadaverine (Ca), and histamine (Hi). The enzymatic activity of DAO towards Hi as a substrate is called histaminase activity12) (Fig. 1). The activity of DAO may be measured in terms of such parameters as the amount of hydrogen peroxide, ammonia, or aldehydes produced in these reactions. Pu or Ca is commonly used to determine DAO activity, but Hi is not.Recently, Sakai et al.13) reported a method to determine Hi using solvent extraction and colorimetry as an associate with Cu(II) and tetrabromophenolphthalein ethyl ester (TBPE) Fig. 1. Scheme of Oxidation of Hi by DAO