A new, highly sensitive and specific method for the fluorometric determination of histamine in human whole blood and plasma is described. The fluorescence spectra and the rate of inactivation by 165-fold purified diamine oxidase from pea seedlings and 120-fold purified histamine methyltransferase from pig antrum mucosa are identical for authentic histamine and the substance isolated from the human plasma. Diamine oxidase was purified by the precipitation of proteins at pH 5.0, ammonium sulphate fractionation at 35 and 65% saturation and by gel filtration on Sephadex G-100. Histamine methyltransferase was purified by ultracentrifugation, ammonium sulphate fractionation at 60 and 80% saturation, chromatography on DEAE cellulose and gel filtration on Sephadex G-. Spermidine did not interfere with the assay of histamine.The normal histamine concentration in human plasma was found to be 0.69 ± 0.26 ng/m/, in human whole blood 54 ± 18 ng/m/. The content of basophil granulocytes in whole blood was shown to be 70 ± 23 cells/mm 3 , whereby more than 90% of histamine in whole blood was localized in basophils. The histamine content of a single basophil granulocyte was calculated to be 1.2 ± 0.4 pg. In a patient, who showed severe anaphylactoid incidents to the anesthetic Propanidid, the plasma contained an average of 7.4 ± 3.2ng histamine/m/. This plasma histamine level is 10-fold higher than the normal values described above. With the aid of the new fluorometric method it seems now possible to investigate the role of histamine in various physiological and pathophysiological processes. Eine hochempfindliche und spezifische Methode zur Bestimmung von Histamin in menschlichem Vollblut und PlasmaZusammenfassung: Eine neue, hochempfindliche und spezifische Methode wird für die fluorometrische Bestimmung von Histamin im menschlichen Vollblut und Plasma beschrieben. Die Fluoreszenzspektren und die Inaktivierungsgeschwindigkeit durch 165fach angereicherte Erbsen-Diamin-Oxidase und 120fach angereicherte Histamin-Methyltransferase aus Schweineantrum waren für Standardhistamin und die aus Plasma isolierte Substanz identisch. Diamin-Oxidase wurde angereichert durch Säurefällung bei pH 5,0, Ammoniumsulfatfraktionierung bei 35 und 65% Sättigung und

ing anaesthesia with both substances led us to undertake this investigation. To demonstrate, however, that the increased concentration of plasma histamine was actually free, pharmacologically active histamine, we studied gastric acid secretion, heart or pulse rate, and peripheral arterial pressure in the subjects of the investigation.

improved method for the determination of histamine release in man: its application in studies with propanidid and thiopentone, European J. Pharmacol. 19 (1972) 180-190. tlistamine release by propanidid and thiopentone was demonstrated with the use of a highly sensitive and specific method for determining histamine in human plasma. 5 min after i.v. administration of propanidid and 3 rain after thiopentone, the plasma histamine level was increased by 350 and 4205~ respectively. In whole blood, histamine release could be demonstrated only when the basophil content remained constant. This was the case in about 50% of test persons after injection of propanidid, but not after administration of thiopentone where the basophil content decreased in all probands by about 35%. The increase of the histamine concentration in plasma was 3 ng/ml and in whole blood, 24 ng/ml, ttalfmaximum gastric acid secretion was elicited by injection of propanidid and thiopentone. It was increased by very rapid application of propanidid and diminished by premedication with atropine by about 30%. The changes in plasma histamine and gastric secretion were parallel over 30 rain, whereas tachycardia and peripheral arterial hypotension lasted for only 3 4 min.During i.v. infusion of histamine a correlation could be shown between the doses infused, (15-90 ng/kg)/min, and the elevation of the plasma histamine concentration. 3 ng/ml plasma were measured at a dose of 145 ng/kg)/min, which induced a half-maximmn gaslric secretion, but was without effect on blood pressure and heart rate. It is concluded that only the increase of plasma histamine concentration, not that of whole blood, represented the release of free, pharmacologically active histamine. In normal test persons, histamine release by propanidid and thiopentone is without significant clinical consequences, but it is important in anaphylactoid incidents occurring during anesthesia with these substances.

Histamine concentrations in canine whole blood and plasma were determined under several pharmacological, pathophysiological, and clinical conditions using fluorometric methods.The specificity of the assay for whole-blood histamine was investigated by comparing 3 purification procedures for the isolation of histamine from whole blood including butanol extraction (Shore), ion-exchange chromatography on Dowex 50 W-X 8, and the combination of these 2 methods (Lorenz). Histamine in whole blood was identified in analytical and preparative samples by fluorescence spectra, thin-layer chromatography, degradation by diamine oxidase from pig kidney and inactivation by histamine methyltransferase from guinea-pig brain as well as by biological tests on the isolated guinea-pig ileum. Since butanol extraction resulted in significantly higher 'histamine' values than the other two purification procedures, ionexchange chromatography on Dowex 50 was recommended as the method of choice for the specific determination of histamine in doffs whole blood.Normal values of histamine concentrations in canine plasma were tentatively estimated. They depended on the time between pretreatment of the animals (anaesthesia, operation) and the collection of blood and showed an approximately logarithmic normal distribution. The median, the lower/upper quartiles and the range of the plasma histamine levels obtained 30 minutes after the end of pretreatment were 0.2, 0-0.4 and 0-1.2 ng/ml, respectively. Nearly 50 % of the values were zero (below 0.1 ng according to the sensitivity of the method), only 1 ~ of them exceeded slightly 1 ng/ml. Thus histamine release by drugs or by other medical treatments was only stated, when plasma histamine levels exceeded 1 ng/ml and decreased in a way to give an elimination carve of approximately first-order kinetics (Bateman function).Histamine concentrations in dog's whole blood showed approximately a logarithmic normal distribution. The median, lower/upper quartiles and range were 47, 34/75 and 13-209 ng/ml respectively.

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