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to infection from the air, especially at the time when sweet, juicy fruits are ripe, when the dust from the ground is very rich not only in yeasts but also in bacteria.
Analyses of air should be performed in breweries from time to time; a clear idea is thus obtained of the progress of the different processes, and will in many cases avert mishap. It is the air of fermenting cellars and coolers in particular to which special attention must be paid.
Soil Analyses. — In soil analyses a small sample is placed in an Erlenmeyer Hask containing a culture liquid chosen with regard to the organisms to be sought. The flora of the soil is a very rich one, especially as regards bacteria and mould fungi. If saccharomycetes are to be looked for, it is advisable to seed the soil sample in wort to which tartaric acid has been added, since this prevents to a great extent the development of most bacteria. In this case also it is desirable to allow the cultures to remain for a considerable time (about fourteen days) at 25° C., and then to prepare a second culture in wort, as saccharomycetes are generally tardy in their development.
It has also been proposed to mix a small quantity of the sample of soil with liquefied nutrient gelatine and then to prepare plate cultures. However, the result is in most cases bad, the number of germs in the soil being too large. Others prefer inaking a paste with the soil in sterile water; plate cultures are then prepared from an average sample.
These analyses, in common with all the foregoing, have the object of discovering the source of the infection which may take place in a brewery. The groundwork for this was furnished by Pasteur's and Hansen's investigations.
11.—Hansen's Pure Culture System in Fermentation Industries.
The Pure Culture System in Bottom Fermentation Breweries. — The following information on the introduction of the systematically selected yeast race into the bottom fermentation brewery is extracted from Hansen's Practical Studies in Fermentation. The starting point must be the yeast which has proved its superiority in working in practice, and has yielded that product which it is wished should be the future normal production of the brewery. Since the species which lends its character in great measure to the product will be present in superior numbers, it will also be the most easily isolated.
If there is wild yeast present in the stock yeast employed, it will, according to Hansen, be present only in small amount in the surface beer at the beginning of the primary fermentation, or it will be totally absent, while at the finish the reverse is the case. A sample of the surface beer is therefore taken just at the time when a frothy head has formed in the fermenting vessel ; we are then certain that the race or species to be isolated is in preponderance, and this sample is used for the preparation of absolutely pure cultures, the starting point being made as usual from a single cell (see page 106). Preliminary fermentations are now made with these pure cultures in flasks in the laboratory. It is advisable to use the same wort as that employed in the brewery, and this should not be re-sterilised in the laboratory. · The method of procuring this is described on page 76. While fermentation proceeds in the flasks, certain preliminary observations are made which will be of use later on. It is observed to what extent the wort remains clear, whether the yeast lies compactly on the bottom, and whether the beer has any peculiar smell and taste, etc. A microscopic examination and also a spore
culture on a gypsum block are of course made; in fact, the characteristics of the yeast are investigated. It may happen that the growths in the flasks which contain pure cultures of the required species are nevertheless somewhat varied, although they belong to the same species. We are here confronted by individual differences, which may always be met with, and we must now make a choice from among these growths also.
After preserving some of the pure culture (on the preservation of yeasts, see page 114) partly in saccharose solution and partly in the dried condition on cotton wool, the procedure is as follows :
Four or five 1 litre Pasteur flasks, containing ordinary aërated but sterilised wort from the brewery, are inoculated from the flask containing the perfectly pure culture; these are set away at the temperature of the room, and in a week they will contain a considerable yeast sediment; four such flasks will generally be sufficient, the fifth being really a reserve culture. After pouring off the beer, the yeast of each of these flasks is introduced into a Carlsberg vessel charged with about 7 litres of brewery wort. After one week as much yeast sediment will have formed in these vessels as is necessary to prepare stock yeast for 1 hectolitre of wort in the brewery. A vessel of 1 hectolitre volume is then set up in the fermenting cellar; this is sterilised by means of a gas flame and charged with a hectolitre of aërated brewery wort. The contents of the four Carlsberg vessels are then poured into this vessel. If the partially fermented wort in the vessels is not to be added also, it is previously drawn off. In the latter case it is advisable to let the flasks stand a little longer, about ten days, so that the yeast may sink more completely to the bottom. If the brewery is some distance from the laboratory it will be always necessary to draw the beer off beforehand. The flasks mentioned above (see page 119) are then used for transporting the thin yeast liquid. The hectolitre of wort mentioned is fermented under the normal conditions of the brewery. If the result is satisfactory the yeast is introduced into practical working.
It is conceivable that a brewery may have operated continuously with a mixture of different species of culture yeast, and that the combination of the latter has formed the character of the product. It is on no account advisable to isolate these species separately and then to employ a mixture of them as stock yeast, for the relative proportions of the species cannot be controlled during fermentation. Besides, it would be far too much trouble in practice.
We have mentioned before that a pure culture yeast of this kind introduced into practice can only keep sufficiently pure in the fermenting vessels for a certain length of time, The resisting powers of the different races against wild yeasts and bacteria is extremely varied. It is, therefore, necessary from time to time to introduce fresh quantities of pure culture yeast into the brewery. There are, however, cases where a race has remained pure in the ordinary fermenting vessels of practice for more than a year without having been renewed.
The Hansen-Kühle Pure Culture Apparatus.-In order to have ready at hand the requisite amount of pure culture yeast, Hansen, in conjunction with Kühle, has constructed a pure culture apparatus for the continuous production in mass of absolutely pure yeast. A description of the apparatus is given below :
The apparatus (Fig. 50) consists of three principal parts, viz., an air pump, A, with an air reservoir, B, a fermentation cylinder, C, and a wort cylinder, D. The air pump receives the air through a filter and pumps it into the reservoir which is provided with a manometer and safety
valve. The air can pass from the reservoir through the communicating pipes, which are provided with outlet taps for condensed water, through the cotton wool filters, g and m respectively, into the fermentation or wort cylinder.
The following parts are connected with the fermentation cylinder, C: (1) a doubly bent side tube, c, which opens under
Fig. 50.-Hansen-Kühle Pure Yeast Culture Apparatus.
water in a vessel, d; (2) a glass tube, f, with marks (10, 20, 79), which indicate the amount of liquid in the cylinder ; taps are fitted at e and h ; (3) a stirring apparatus, b, to stir up and distribute any bottom yeast formed in the cylinder ; (4) an outlet cock, 1, for beer and yeast ; (5) a short side tube, j, provided with rubber tubing and glass stopper; the pure