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1. A simplification of the method of Anrep et al. for the estimation of histamine in human urine is described. 2. It is not possible by this method to obtain a reliable estimation of the amount of free histamine normally present in human urine, but the effects of the administration of histamine may be followed. 3. When 133 mg. of histamine was given by mouth to three men, it was mainly conjugated histamine that appeared in the urine. 4. When 3·5‐5 mg. was infused intravenously in three men, free histamine appeared in the urine. 5. In two of these experiments no changes were detected in the histamine content of the plasma or cells obtained from the antecubital vein. It is concluded that investigations of the excretion of histamine are more likely to give information about the release of histamine in the body than investigations on venous blood.

The method is intended for the estimation of minute quantities of histamine in blood plasma. A neutralized trichloracetic acid extract of plasma is adsorbed at pH 8 on a column prepared by mixing a quantity of the cationic exchange resin Amberlite XE-64 with powdered cellulose as a supporting medium. Histamine is adsorbed but N-acetylhistamine is not: the amines can therefore be estimated independently. Elution of the histamine is by displacement with HCL. The eluate is converted into the solution for bioassay on the superfused guinea-pig ileum.In recovery experiments histamine was added in the range 25 to 100 ng. to solutions of known composition and to plasma (5 ml.) obtained from man and the cat. The mean recovery in this range for histamine added to plasma was 82.5% A2 S.E. (17 estimations).The histamine equivalent of human plasma obtained from the antecubital vein was found to be less than 1 ng./ml.It was possible by this method to follow the concentration of histamine in the arterial plasma of the cat when histamine was infused intravenously at a rate of 330 ng./kg./min. There was no perceptible change in the arterial blood pressure during the infusion, but the plasma histamine rose from <0.3 ng./ml. to 3.4 ng./ml. (mean of 2 estimations).When the quantity of histamine extractable from plasma is less than can be estimated by conventional methods of bioassay, the sensitivity of the test can be increased by applying the undiluted extract directly to the strip of guinea-pig ileum; this may be suspended in a minute bath (Mongar and Schild, 1950) or in air, as in the technique of superfusion (Gaddum, 1953). The method is applicable if the histamine has been sufficiently purified and if the test solution is similar in composition to the solution bathing the intestine between the doses.If plasma is extracted by the method of Barsoum and Gaddum (1935) as modified by Code (1937a) and tested in this way, the activity of the extract is only partially abolished by the histamine antagonist mepyramine maleate (Mongar and Whelan, 1953). Moreover, the method does not distinguish between free histamine and histamine which may be released from pharmacologically inactive forms at the stage of refluxing the extract in strong acid (unpublished results).Various chromatographic methods of purifying histamine have been proposed and are reviewed in the article by Code and McIntire (1956). Some employ paper chromatography, others weak cationic exchangers which are used to remove histamine from solutions containing organic solvents.These methods are designed mainly for the chemical determination of histamine, but some have been combined with bioassay.In the method to be described the purification is conducted entirely in aqueous solution with the aid of a carboxylic ion exchange resin, at room temperature and at pH values not far from neutral. Advantage is taken of the fact that carboxylic resins have a high exchange capacity and can be buffered over a wide range of pH. They can therefore be used to separate weak organic bases in a...

The effect of intravenous infusions of histamine on gastric secretion has recently been studied quantitatively in the cat (Wood, 1948) and in the dog (Obrink, 1948;Hanson, Grossman, and Ivy, 1948). These authors observed that the effect of the drug increased with the dose, and that by choosing a suitable range of doses it was possible to obtain data which could be used to construct doseresponse curves.Little is known about the effect of histamine infusions on gastric secretion in man. According to McElin and Horton (1946) ng./kg./min. Our aim was to study the secretory behaviour of the human stomach in response to prolonged continuous intravenous infusions of histamine, and to determine the relationship between dose and response. At the same time we attempted to gain further information on the fate of histamine in the body by estimating it in samples of various body fluids, including the gastric juice. This part of the work has been reported elsewhere (Adam, Card, Riddell, Roberts, and Strong, 1954).Our data were obtained from three healthy men designated as W.I.C., J.A.S., and A.A.G., each of whom received a series of intravenous infusions of histamine. Estimates of the rates of acid secretion were plotted against rates of infusion of the drug. Each infusion was thus a self-contained experiment and could be represented by a point on a graph. By applying statistical methods to the results it was possible to fit various parameters of the doseresponse curves. METHODS Intubation of the Stomach and Collection of the Juice. -The subject omitted breakfast. A few drops of local anaesthetic (Amethocaine HCI) were instilled into the nose and a Ryle's.tube was passed by this route into the stomach. The position of the tip was ascertained radioscopically and the tube manipulated until the tip lay on the left border of the vertebral column. The subject then lay on his left side, and the stomach contents were aspirated by syringe. This was sometimes made easier by washing the stomach out with warm water. The Ik yle's tube was then connected to a source of continuous suction not exceeding 40 mm. Hg. The gastric juice flowed into a 50-ml. boiling tube, which was replaced every ten minutes by a clean tube. At slow rates of secretion the juice tended to be thick and mucinous and occasionally blocked the tube. The injection of a small quantity of warm water easily cleared the tube, and a correction was subsequently applied to the volume of the sample. saliva was collected by expectoration. The duodenal juice was not withdrawn.Titrations.-The volume of the sample of gastric juice was measured and the free and total acid were determined by titration against 0.lN-NaOH, using 'I pfer's reagent and phenolphthalein as internal indicators. The concentration of acid was expressed in m.equiv. HCl/l. The product of the concentration of total acid and the volume of the sample in litres gave an estimate of the output of acid in m.equiv. HCI.Infisions.-Histamine acid phosphate was supplied by British Drug Houses Ltd. All the values for histam...

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Chromatographic purification, combined with very sensitive biological methods of assay, can greatly facilitate the quantitative estimation of physiologically active compounds occurring at extremely low concentrations in body fluids. New techniques of this kind appear to be essential before further progress can be made in the investigation of any role histamine might play in normal human physiology. Free, pharmacologically active histamine has not yet been reliably estimated in human plasma. Human urine has been shown to contain fair quantities of histamine, but almost entirely in a conjugated form (Anrep, Ayadi, Barsoum, Smith, and Talaat, 1944). Urbach's work (1949) suggests that urinary conjugated histamine is acetylhistamine [4(3-acetylaminoethyl)imidazole], and Tabor and Mosettig (1949) were able to isolate this compound from the urine of dogs fed with histamine and to identify it chemically. Evidence for the normal appearance of free histamine in human urine is not so clear-cut. Ackermann and Fuchs (1939) prepared from a single sample of 10 litres of urine, by adsorption on Lloyd's Reagent and purification through the phosphotungstate, a quantity of histamine the bioassay of which implied a concentration of 11.7 vg./litre of free base in the original urine. But Anrep et al. (1944), using a charcoal-adsorption method, were able to detect no more than traces of free histamine in normal human urine, a result confirmed later by Adam (1950).In attempting to work out simpler and more sensitive methods, we have examined a number of chromatographic materials for their capacity to adsorb histamine and acetylhistamine. Adsorbents able to take up these substances directly from body fluids were specially looked for. Such materials would make easier the concentration of these compounds (free histamine in particular) from relatively large volumes of fluid. An adsorbent which could separate histamine and acetylhistamine quantitatively would be extremely valuable, and would make possible the independent determination of these substances in fluids where they occur together. As the quantities expected are minute, sensitive and relatively specific biological methods of assay have been used in preference to chemical methods.The adsorption of histamine and its separation from acetyihistamine Permutit-like materials have been tried out for histamine by Whitehorn (1923), Schwartz andRiegert (1936), andCode andIng (1937). Such adsorbents act by cation-exchange, and are capable of taking up the ions of organic bases from very

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