The estimation of catecholamines in urine by a chemical method.

The estimation of urinary catecholamines has assumed clinical significance mainly because of its value in the diagnosis of phaeochromocytoma (Engel and Euler, 1950; Euler, 1951). Since biological methods have obvious limitations for routine work, various chemical methods have been developed (Lund, 1950; Pekkarinen and Pitkanen, 1955; Euler and Floding, 1955a, b). They are based on the same principle: catechol compounds are isolated by adsorption on aluminium oxide and adrenaline and noradrenaline are subsequently oxidized to fluorescent trihydroxyindoles. In addition to adrenaline and noradrenaline a third catecholamine occurs in urine, namely 3-hydroxytyramine (Holtz, Credner, and Koepp, 1942; Euler and Hellner, 1951 ; Euler, Hamberg, and Hellner, 1951). Few quantitative data about its rate of excretion are available at present (Euler et al., 1951; Hoobler, Agrest, and Warzynski, 1954; Abelin and Goldstein, 1955); they were obtained by means of chromatographic procedures which can, at best, be regarded as semi-quantitative. In the method here proposed the three catecholamines are quantitatively estimated by a combination of the fluorimetric methods of WeilMalherbe and Bone (1952) and Euler and Floding (1955a). The former estimates the sum of the three catecholamines; adrenaline and noradrenaline are estimated separately by the second method; hydroxytyramine is obtained by difference. Euler, Euler, and Floding (1955) have shown the occurrence of re'latively large amounts of 3:4dihydroxyphenylacetic acid (catecholacetic acid) in urine. This substance and any other acidic catechol may be separated from catechol bases by ion exchange. After passage through an anion exchange resin the catechol bases are found in the filtrate. On a cation exchange resin they are themselves retained. We have chosen cation exchange as the more specific procedure in which neutral interfering substances whose presence in urine has been claimed, such as catechol (Fiker, 1953) or 3: 4-dihydroxybenzaldehyde (Kawamoto, 1954), would also be eliminated. The resin treatment is preceded by the passage of the urine sample through a column of aluminium oxide, essentially as described for plasma (Weil-Malherbe and Bone, 1952). The higher purification resulting from the cation exchange procedure is of advantage even if it is desired to estimate only adrenaline and noradrenaline. Better results were obtained when the methods of Lund (1950) or of Euler and Floding (1955a) were applied to urine extracts purified in this way than when they were used in their original form, i.e., after alumina adsorption only. The advantages were that the blank was lower, indicating the removal of non-specific fluorescence, and that recoveries were better, indicating perhaps the removal of quenching impurities. Eluates from alumina columns usually contain a reddish pigment which forms a flocculent precipitate in alkaline solution. This material is eliminated in the resin treatment.