The assay of vitamin B4

Abstract

IN earlier publications [Reader, 1929, 1930] evidence was presented for the division of the vitamin B complex into three components all necessary for the nutrition of the rat'. It was shown that two of these (Bl and B4) were alkali-labile while B2 was not appreciably destroyed by autoclaving a crude yeast extract solution at PH 9 for one hour at 1200 [see Reader, 1930, p. 79]. The work reported here deals with the methods of assay used during the concentration and purification of the second alkali-labile factor B4. In the earlier papers the restoration of growth of young rats, which had come to constant weight on a basal diet [Reader, 1929] plus excess of vitamlins A, B, B2 and D, was used as a criterion of the presence or absence of vitamin B4 in a given extract. As there is a prolonged storage' period for vitamin B4 a certain amount of difficulty was experienced in keeping the animal colony sufficiently deficient in this vitamin, while, at the same time, supplying essential natural products (milk, etc.) to the breeding rats to get adequate lactation. Consequently a new "adult-curative" method was devised. Use of this method has led to the surprising discovery that the symptoms usually considered to be typical of polyneuritis in rats are really a mixture of symptoms due to lack of vitamins B, and B4. By eliminating the vitamin B, deficiency at the critical stage the true clinical picture of vitamin B4 deficiency is revealed and can be used in a curative test. The main features of the adult-curative test are as follows. The animals, grown to maximum weight with adequate, but not excess, vitamin B4 for normal growth, were completely deprived of both the alkali-labile vitamins B, and B4 until polyneuritis occurred (3 to 4 weeks). At this stage it was found that vitamin B, cures the typical polyneuritis symptoms [see Kinnersley, Peters and Reader, 1930] but it does not restore weight or ameliorate all the symptoms, even when given in large excess doses daily, up to 12 pigeon doses per day. Instead the animal remains in a weak condition with swollen red' paws, spastic gait, loss of coordination , and other symptoms apparently due to the specific lack of vitamin B4. Now if a positive test dose of vitamin B4, proved free from vitamin B,, 1 The term B4 is used throughout this paper to denote the third rat "B" factor, and is equivalent to B. in the earlier papers by the author. Thus it is hoped to avoid confusion with the Williams and Waterman thermolabile pigeon factor which has been termed B3.