PREVIOUS papers from this laboratory have dealt with the application of the titration method with 2: 6-dichlorophenolindophenol to the estimation of vitamin C in various animal materials [Harris and Ray, 1933, 1, 2; Birch et al., 1933; Birch and Dann, 1933; Harris, 1933]. Results on human urines are discussed in the present communication. A principal purpose we had in mind in undertaking this enquiry was to discover whether any relation could be found between urinary loss and the state of nutrition of the individual in respect to vitamin C sufficiency or subnormality. Any such index would obviously be of great value for practical human dietetics.Previous biological work has failed to demonstrate the presence of vitamin C in urine [van der Walle, 1922], but a note just published by Eekelen et al. [1933] mentions, without giving actual data, that the reducing substance in urine is higher in persons using much fruit. METHOD.The micro-method of Birch et al. [1933] was used throughout. The urine to be examined was made acid with trichloroacetic acid (final concentration of latter, 5 %) and titrated from a micro-burette reading to 0 01 cc. against 005 cc. of the indicator, which had been previously standardised against ascorbic acid. Titrations were carried out immediately after, or within a few minutes of, urination, as we found that the titre tended to fall in urine which had been allowed to stand for long. It is essential to carry out the titration rapidly and reach the end-point within about 1 min., otherwise erroneously high values will be caused by phenolic or similar substances in the urine reducing. the indicator slowly. Urines are sometimes encountered which are too dilute to give a satisfactory end-point. If a reading is required for such an individual it is necessary to restrict his fluid intake. The results to be described make it clear that the titration figure bears a genuine relation to vitamin C metabolism, but in expressing our results for convenience in terms of so many mg. of ascorbic acid we wish to. make it clear that it is without prejudice to the question of the invariable specificity or otherwise of the reaction. EXPERIMENTAL.Excretion after large dose. Our most detailed observations were undertaken with the object of obtaining a quantitative picture of the course of excretion of the vitamin after the administration of a single large dose. These results will be

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Until abouit 8 years ago the only method for determining vitamin C was by means of biological tests on guinea-pigs. In consequence, knowledge about the distribution and behaviour of the vitamin accumulated slowly and with difficulty, and it was not until a chemical method had been introduced that progress became more rapid.

IN earlier work [Harris & Leong, 1936; Harris et al. 1938] it was shown that the extent of the body's "reserves" of vitamin B1 might be measured by means of a test on the urine, similar in principle to that previously introduced for vitamin C [Harris et al. 1933; Harris & Ray, 1935]. The urinary output was found to depend on the past intake, being negligible in beri-beri or in "conditioned" deficiencies of vitamin B1 and being lowered in certain other abnormal states.To simplify the procedure, several investigators [e.g.

It has often been assumed that animals given a suitable free choice of diets are able to select satisfactorily according to their individual nutritional needs. However, no systematic work has hitherto been undertaken to ascertain how far this is true, nor have we any knowledge of the mechanism involved in the choice. Lusk (1928), in his well-known text book on nutrition, refers to the observation of a German worker (Tscherkes, 1923) that fowls suffering from polyneuritis will search out green food and refuse to accept grain. This be describes as due to the "triumph of instinct." The similar eagerness of vitamin B deficient rats to consume diets containing the vitamin must, we imagine, have been noticed by observant workers. It is, however, unsatisfactory to allow the matter to be ascribed vaguely to some unexplained—it nor inexplicable—" instinct." It animals are in tact able to distinguish between foods containing and deficient in the vitamin, what is the nature of the process involved? Has the animal some means, as by taste or smell, of recognizing the vitamin per se ? It so, it would imply an ability to detect a constituent amounting to no more than perhaps 1 part in a million of the food. It nor, by what means is the rat able to recognize the vitamin-containing food ? And what are the limitations to such powers of recognition ? These are some of the questions which one sought to answer, An effort to analyse these phenomena was begun in this laboratory in 1928, and in the present paper are summarized the principal results reached during the period 1928-1931.* To anticipate a main conclusion it max be said at once that we have obtained good evidence that the behaviour of the animal is due not so much to instinct as to experience, i. e ., of the beneficial effect produced by a particular foodstuff. We believe that This factor of experience plays an important part in determining dietary preferences in general, and it will certainly have to be taken into account in future work on that hitherto neglected subject, the psychology of appetite. Apart from the reference quoted above we know of no previous literature relating to the original point of departure of our enquiries. Recorded work on the whole question of the free choice of diet is almost equally scanty. Osborne and Mendel (1918), working on the comparative food values of different proteins, thought that rats " as a rule ate more of the adequate than of the inferior food " ; but Beadles, Braman and Mitchell (1930), on the contrary, could find " no support for the assumption that the more complete of the two rations is consumed in greater amount." Nevens (1928) noted that cattle given a free choice of limestone, bone meal and salt, offered as supplements to an inadequate diet, took only insignificant amounts of the first two ; while kon (1931) found that rats might fail to take enough protein even to keep them alive, when allowed to choose their own allowance of carbohydrate, protein and salt mixture.

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IT is already known that the suprarenal gland is an extremely potent source of vitamin C, but hitherto the activity has been thought to be restricted to the cortex. In the present paper, however, it is shown that the medulla of the suprarenal is also intensely active, its potency falling but little short of that of the cortex. Our results indicate that ox suprarenal cortex has an activity of about 30 international units per gram and ox medulla of about 20 units, i.e. no less than two-thirds of the former. In other words the cortex has about thrice and the medulla about twice the potency of fresh orange juice or lemon juice (the international standard of vitamin C activity). Szent-Gy6rgyi [1928], struck by the peculiar silver-reducing power of the cortex, isolated from it the substance thought to be responsible-first called hexuronic acid, but later, after its identification with vitamin C, renamed ascorbic acid. The intense antiscorbutic activity of the suprarenal cortex was demonstrated in papers from this laboratory [Harris, Mills and Innes, 1932; Harris and Ray, 1932; 1933, 1] and the degree of activity was shown to be commensurate with its richness in hexuronic acid. A chemical method for estimating the latter, based on modifications in the use of Tillmans's oxidationreduction indicator 2: 6-dichlorophenolindophenol was worked out and applied to various animal and vegetable materials, and the surprising discovery was made that certain tissues, including the suprarenal medulla, not hitherto recognised as sources of vitamin C, also gave a high titration value [Harris and Ray, 1933, 1, 2; Birch, Harris and Ray, 1933; Birch and Dann, 1933; Harris, 1933, 1]. This result at once suggested that the absence of the silver reaction was in reality little guide as to the presence or absence of vitamin C, a conviction which was strengthened by our further observation that the medulla and cortex of different species, as well as a variety of other tissues (e.g. liver), might stain or not in the most erratic order with little apparent parallelism with their true antiscorbutic activities. Again liver extract and other tissue extracts although rich in vitamin C did not reduce silver. Similarly Gough and Zilva [1933] have recently noted that human suprarenals may fail to darken at all with silver (their vitamin C potency, however, not having been tested). The chemical test for vitamin C, although it is known to be reasonably specific, may give high results in certain exceptional cases, and it was therefore necessary to check the titration results on the suprarenal medulla against direct feeding tests. The biological determination gave results in complete agreement with the indophenol titration. We have recently repeated these feeding tests and exactly confirmed our earlier results.