Fig. 29.—Bran Fermentation. Advanced Stage.
A fairly pure culture may be obtained by inoculating a tube of bran infusion, previously sterilized, by means of a freshly made capillary pipette, which is pushed into the tube without removing the cotton-wool plug. The tube is placed in the incubator at 30° C., and the organisms causing the fermentation develop rapidly; as soon as the tube is observed to become cloudy, indicating the development of the bacteria, another tube is inoculated in the same manner, using a very minute quantity of liquid; this second “attenuation” is then carried on to five or six more tubes. In the fifth tube will usually be found a pure culture of the bran fermenting organisms, which I have called Bacterium furfuris. The bacteria (see Fig. [30]) are mostly in the form of dumb-bells or pairs, each cell 0·75 µ × 1·25 µ: they vary slightly in size, some forming chains; those on the surface become surrounded by a kind of jelly, and form an iridescent pellicle on the surface of the liquid, the zoogloae form. When the nutriment of the liquid is exhausted or the acidity becomes too great,[154] this film sinks to the bottom and the liquid above becomes comparatively clear. There is no spore formation, and, consequently, the culture soon dies unless inoculated into a fresh nutrient medium. The liquor, after the fermentation both in the drenches and tubes of pure cultivations, is always slightly ropy.
B. furfuris probably belongs to the group of coli bacteria, some of which ferment milk and other sugars with production of lactic acid and gases. This explains the ease with which the fermentation takes place, since the skins bring in immense numbers of bacteria from the bate, among which are the organisms described.
Eitner[155] states that the action of the bran drench cannot be considered a chemical one, and that the chief action is a dynamical one, resulting from the gases generated in the liquid and in the skin. While I do not deny that the gas has such action—indeed I pointed out in 1893[156] that the gas acted by floating and distending the skins, and so enabling them better to take up the acids—I maintain, however, that the chief action is a chemical one and due to the weak organic acids produced; this was confirmed experimentally. The researches on bran drenching commenced in 1887 by myself, and later in conjunction with Dr. W. H. Willcox, prove conclusively that the amount of acid produced, and especially the lactic acid, is sufficient to produce a very decided action on skin. In actual working we found from 1·07 to 2·34 grm. total acids per 1000 c.c., while in artificial cultures, with addition of calcium carbonate, inoculated with pure cultures of bacteria, obtained from drenches, we obtained from 4·53 to 11·44 grm. total acids per 1000 c.c. Now traces of lime in the skins act in just the same way as the calcium carbonate; they neutralize a portion of the acid produced by the bacteria, and so allow a greater amount of acid to be produced than if it accumulated in the liquid. By preparing solutions of the above acids of suitable strength and agitating skins in them, results were obtained in 1 1/2 hours equal to a drenching of 14 hours. The defect of such artificially prepared liquids in practice is due to the fact that they do not contain any particles of bran or flour; these take up dirt from the skin mechanically, and so produce a better-coloured leather. Where the skin has been bated in a perfectly clean bate, and then paddled in such an artificial drench, the leather is in every way equal to that prepared by means of the dung bate, followed by a bran drench.
By far the most important action for the tanner is the solution of the last traces of lime not dissolved by the bate, and this is the chief reason for its use, where colour is a desideratum, and where sweet liquors are used; for some kinds of leather where it is desirable not to lose pelt, the drench is used alone for this purpose. A dung bate, such as most light leather tanners use, is seldom run off more than once a week, and contains a considerable amount of dissolved lime; the amount in a freshly made-up puer wheel is from 0·1 grm. to 0·3 grm. lime (CaO) per litre. After the wheel has been in use for a week, it rises to 0·5 grm. to 0·8 grm. per litre. It will be readily seen that a liquid which already contains such considerable quantities of lime is not an especially good medium to use for its complete removal from the skins, hence drenching is necessary to get rid of the lime left in the skins.
Eitner states that the fermentation is alcoholic at the commencement, and then becomes an acid fermentation with generation of gas. In tan liquors there is no doubt that this is the course of the acetic fermentation, as has been shown by Andreasch,[157] but in the usual type of bran drench the action is certainly a different one. The acetic acid, as far as we can ascertain, is produced directly from the dextrose, without the previous production of alcohol, since the presence of the latter is not shown by tests in any stage of the fermentation.[158] No yeasts were observed[159] and no alcohol was found in any stage of the fermentation.[160] J. O’Sullivan remarks that although it has not been noticed it is possible that in this case the production of alcohol and its oxidation into acetic acid are simultaneous, but that there is no preliminary fermentation by yeasts followed by acetic fermentation by bacteria such as Eitner indicates is certain.
To sum up the conclusions arrived at from the experiments above quoted.