Attempts to prepare active extracts from undried yeast in an analogous manner have so far not been very successful. Thus Rinckleben [[1911]] found that plasmolysis by glycerol (8 per cent.) or sodium phosphate (5 per cent.) sometimes yielded an active juice and sometimes a juice which contained enzyme but no co-enzyme, but more often an inactive juice incapable of activation (p. [64]) [see also Kayser, [1911]].
Giglioli [[1911]] by the addition of chloroform also obtained an active liquid. It appears in fact as though almost any method of plasmolysing the yeast cell may yield a certain proportion of zymase in the exudate.
An ingenious process has been devised by Dixon and Atkins [[1913]] who applied the method of freezing in liquid air which they had found efficacious for obtaining the sap from various plant organs. They thus succeeded in obtaining from yeast, derived from Guinness' brewery in Dublin, liquids capable of fermenting sugar and of about the same efficacy as the maceration extracts prepared by Lebedeff's method from the same yeast. The results were, however, in both cases very low, the maximum total production of CO2 by 25 c.c. of liquid from excess of sugar being 32·5 c.c. (air temperature) or about 0·06g. Munich yeast on the other hand yields, either by maceration or grinding, a liquid giving as much as 1·5–2g. of CO2 per 25 c.c., whilst [p027] English yeast-juice prepared by grinding often gives as much as 0·5–0·7g. of CO2.
No direct comparison with the juice prepared by grinding was made by Dixon and Atkins, but it may be concluded from their results that the best method of obtaining an active preparation from the top yeasts used in this country is that of grinding. Maceration, freezing and plasmolysis alike yield poor results. With Munich yeast on the other hand the maceration process yields excellent results, whilst the liquid air process has not so far been tried.
Practical Methods for the Estimation of the Fermenting Power of Yeast-Juice.
In order to estimate the amount of carbon dioxide evolved in a given time and the total amount evolved by the action of yeast-juice on sugar, Buchner adopted an extremely simple method, which consisted in carrying out the fermentation in an Erlenmeyer flask provided with a small wash-bottle, which contained sulphuric acid and was closed by a Bunsen valve, and ascertaining the loss of weight during the experiment. Corrections are necessary for the carbon dioxide present in the original juice and retained in the liquid at the close of the experiment and for that present in the air space of the apparatus, but it was found that for most purposes these could be neglected. In cases in which greater accuracy was desired, the carbon dioxide was displaced by air before the weighings were made. A typical experiment of this kind, without displacement of carbon dioxide, is the following:—
| March 22, 1899, Berlin bottom yeast V. 20 c.c. juice + 8 grams cane sugar + 0·2 c.c. toluene as antiseptic at 16°. Grams of carbon dioxide after | ||||
|---|---|---|---|---|
| 24 | 48 | 72 | 96 hours. | |
| 0·40 | 0·64 | 0·99 | 1·11 | |
The total weight of carbon dioxide evolved under these conditions is termed the fermenting power of the juice (Buchner).
A more accurate method [Macfadyen, Morris, and Rowland, [1900]] consists in passing the carbon dioxide into caustic soda solution and estimating it by titration. The yeast-juice, sugar, and antiseptic are placed in an Erlenmeyer flask provided with a straight glass tube, through which air can be passed over the surface of the liquid, and a conducting tube leading into a second flask which contains 50 c.c. of 10 per cent. caustic soda solution and is connected with the air by a guard tube containing soda lime. The juice can be freed from carbon dioxide by agitation in a current of air before the flask is connected to [p028] that containing the caustic soda solution, and at the end of the period of incubation air is passed through the apparatus, the liquid being boiled out if great accuracy is required. The absorption flask is then disconnected and the amount of absorbed carbon dioxide estimated by titration. This is carried out by making up the contents of the flask to 200 c.c., taking out an aliquot portion, rendering this exactly neutral to phenophthalein by the addition first of normal and finally of decinormal acid, adding methyl orange and titrating with decinormal acid to exact neutrality. Each c.c. of decinormal acid used in this last titration represents 0·0044 gram of carbon dioxide in the quantity of solution titrated.
