The formaldehyde-azo-test for vitamin B1

Abstract

THERE is some vagueness as to the exact scope of the so-called Pauly reaction, such as is given by histidine with diazotised sulphanilic acid. In the broad sense, it might be considered to characterise any substance reacting in a weak alkaline solution to give the characteristic pink or red colour. In the particular sense however it is often now taken to indicate a substance reacting with the reagents studied in detail by Koessler and Hanke [1919; see also Hunter, 1922;1925].Jansen and Donath [1926] Hence we were correct in attributing our previous 3 % to impurities.Nevertheless, in essence the opinion as expressed at the British Association Meeting is correct, namely that the vitamin would interact with the diazoreagent to give a pink colour at a more alkaline reaction than the one employed in the Koessler and Hanke method. We have delayed publication on this question for over two years in the effort to collect conclusive evidence, and we are now presenting shortly some of our data (as a preliminary step). Two circumstances have assisted our progress: (1) the realisation that rather fine shades of alkalinity 1 It should have been added "in this sense."

Improvements in the use of the formaldehyde azo reaction for vitamin B1

“…Nevertheless, we think that some independent check may prove useful, and further, that the method may prove of value, when enough vitamin B1 is available, owing to the stability of the standards, and to the fact that a dark room and a source of ultra-violet light are not required. Unfortunately, neither method is successful with phosphoric esters of vitamin B1; these do not give the formaldehyde azo test (as stated by Lohmann & Schuster [1937]); they form a blue fluorescent compound with alkaline ferricyanide, but this does not pass into a butyl alcohol layer, which is a basis of separation from interfering fluorescent substances [Kinnersley & Peters, 1937]. We have found that phosphatase can be used inthe pre-treatment of the extracts to liberate the vitamin B1.…”

“…After further cooling and removal of insoluble matter by the centrifuge, Na phosphotungstate, 10 % at pH 6, is added to slight excess and the mixture is acidified to pH 1 0 with 20% H2S04. The phosphotungstate precipitate is allowed to stand 12 hr., collected in the centrifuge tube, and ground with baryta three times, according to our usual technique [Kinnersley et al 1935]. After removal of baryta by H2S04, the extract is concentrated if necessary at pH 3, care being taken to see that greater acidity does not develop during concentration; any free H2S04 should be removed by BaCl2 during this process.…”

“…THIs paper embodies further observations made upon and in relation to the formaldehyde azo reaction for vitamin B1 [Kinnersley & Peters, 1934] and its' possible application to semi-quantitative estimation of vitamin B1. The development of the more sensitive thiochrome method of estimation [Jansen, 1936;Karrer & Kubli, 1937;Pyke, 1938] during the progress of this work, may ultimately make the method here described obsolete.…”

“…The question arises whether the presence of a metal could explain the delayed azo reaction previously reported by Kinnersley et al [1935] during the formation of quinochromes by oxidation with manganese oxides. If Mn compounds could produce the effect this would be reasonable; but they do not do so, nor is it possible to modify the reaction for vitamin B1 by addition of the solution in which the vitamin is oxidising.…”

über den Vitaminhaushalt in der Schwangerschaft und im Wochenbett bezüglich der A‐ und B1‐ Vitamine, der Nikotinsäure und des C‐ Vitamins

“…Von den chemischen Bestimmungsverfahren des B,-Vitamins sind einige kolorimetrisch und beruhen darauf, dass gewisse chemische Substanzen mit B,-Vitamin eine Farbreaktion geben. Zu diesen gehort zunachst der Formaldehyd-azo-Test von KINNERSLEY und PETERS (1934), wonacli das B,-Vitamin in alkalischer Losung mit Diazobenzolsulfosaure unter Bildung von Rotfarbung reagiert. Nach der Methode von PRC-B L U D A und MCCOLLUM (1936) wiederum gibt p-Amino-azetanilid und p-Amino-Azetophenon rnit B,-Vitamin in Wasser schwerlosliche rote Verbindungen, und WILLSTAEDT und BARANY (1938) benutzten zur Bestimmung des Aneurins 2,4-Dichlorbenzoldiazoniumchlorid, welches rnit B,-Vitamin in alkalischer Losung unter Bildung von Azofarbstoff reagiert .…”