Alcoholic fermentation / by Arthur Harden.

  • Arthur Harden
Date:
1932
Catalogue details

Licence: In copyright

Credit: Alcoholic fermentation / by Arthur Harden. Source: Wellcome Collection.

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    Occasionally the precipitate of lead salt retained some of the co-enzyme, apparently by adsorption, but usually the greater part remained in solution. It was also known that the co-enzyme is partially removed from yeast-juice by means of a colloidal solution of ferric hydroxide (Resenscheck). A precipitate is thus obtained which contains phos- phorus and resembles boiled yeast-juice in its regenerative action on yeast-juice rendered inactive by fermentation. It has not, however, so far been found possible to isolate any definite compound from this precipitate. There are also indications that when yeast-juice, either fresh or boiled, is electrolysed, the co-enzyme tends to accumulate at the cathode [Resenscheck, 1908, I, 2]. Euler and Myrback [1924, 2, 4; 1928, I; 1929; Myrback and Euler, 1924] start with a yeast extract made by heating yeast with 5-6 parts of water at 90°, and evaporating at a low temperature. This is first dialysed against water in collodion membranes and the dialysate, which is free from colloidal matter, is used. This is treated with lead acetate at pn 6-0 and the filtrate, which contains the co-enzyme, is precipitated with lead acetate and sodium hydroxide at pK io-o. The washed precipitate, containing the co-enzyme, is decomposed by sulphuric acid and from the resulting solution the co-enzyme is pre- cipitated by addition of mercuric nitrate, sufficient sodium hydroxide being added to keep the liquid only slightly acid. The well-washed precipitate is then decomposed by H2S and the latter removed by a current of air. The double precipitation can be avoided by adding the mercuric nitrate directly to the filtrate from the neutral lead acetate precipitation, but the product is not so pure. Further purification is effected by adding silver nitrate to the acid solution resulting from the decomposition of the mercuric nitrate precipitate, filtering and to the filtrate adding excess of silver nitrate and then just sufficient dilute ammonia to produce maximum precipita- tion. The precipitate is washed with water and decomposed by H2S. This solution (ACo = 20,000) is precipitated with phosphotung- stic acid in 2 per cent, sulphuric acid, the precipitate washed with dilute sulphuric acid and extracted with ether and amyl alcohol in presence of very dilute sulphuric acid to remove the phosphotungstic acid, and from this solution the co-enzyme is precipitated, after con- centration in vacuo, by the addition of a large volume of alcohol or acetone. A perfectly white preparation is obtained of high activity (ACo = 30-50,000). This material is not an individual substance since picric acid precipitates from its concentrated solution the picrate of adenylthiomethylpentose, CnH1503N5S, the presence of which in
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