Exactly half of the lime in the tartrate employed got used up in the soluble salts formed during fermentation; the other half was partly precipitated in the form of carbonate of lime, partly dissolved in the liquid by the carbonic acid. The soluble salts seemed to us to be a mixture or combination of 1 equivalent of metacetate of lime, with 2 equivalents of the acetate, for every 10 equivalents of carbonic acid produced, the whole corresponding to the fermentation of 3 equivalents of neutral tartrate of lime. [Footnote: The following is a curious consequence of these numbers and of the nature of the products of this fermentation. The carbonic acid liberated being quite pure, especially when the liquid has been boiled to expel all air from the flask, and capable of perfect solution, it follows that the volume of liquid being sufficient and the weight of tartrate suitably chosen—we may set aside tartrate of lime in an insoluble, crystalline powder, alone with phosphates at the bottom of a closed vessel full of water, and find soon afterwards in their place carbonate of lime, and in the liquid soluble salts of lime, with a mass of organic matter at the bottom, without any liberation of gas or appearance of fermentation ever taking place, except as far as the vital action and transformation in the tartrate are concerned. It is easy to calculate that a vessel or flask of five litres (rather more than a gallon) would be large enough for the accomplishment of this remarkable and singularly quiet transformation, in the case of 50 grammes (767 grains) of tartrate of lime.]. This point, however, is worthy of being studied with greater care: the present statement of the nature of the products formed is given with all reserve. For our point, indeed, the matter is of little importance, since the equation of the fermentation does not concern us.
After the completion of fermentation there was not a trace of tartrate of lime remaining at the bottom of the vessel: it had disappeared gradually as it got broken up into the different products of fermentation, and its place was taken by some crystallized carbonate of lime—the excess, namely, which had been unable to dissolve by the action of the carbonic acid. Associated, moreover, with this carbonate of lime there was a quantity of some kind of animal matter, which, under the microscope, appeared to be composed of masses of granules mixed with very fine filaments of varying lengths, studded with minute dots, and presenting all the characteristics of a nitrogenous organic substance. [Footnote: We treated the whole deposit with dilute hydrochloric acid, which dissolved the carbonate of lime and the insoluble phosphates of calcium and magnesium; afterwards filtering the liquid through a weighed filter paper. Dried at 100 degrees C. (212 degrees F.), the weight of the organic matter thus obtained was 0.54 gramme (8.3 grains), which was rather more than 1/200 of the weight of fermentable matter.] That this was really the ferment is evident enough from all that we have already said. To convince ourselves more thoroughly of the fact, and at the same time to enable us to observe the mode of activity of the organism, we instituted the following supplementary observation. Side by side with the experiment just described, we conducted a similar one, which we intermitted after the fermentation was somewhat advanced, and about half of the tartrate dissolved. Breaking off with a file the exit-tube at the point where the neck began to narrow off, we took some of the deposit from the bottom by means of a long straight piece of tubing, in order to bring it under microscopical examination. We found it to consist of a host of long filaments of extreme tenuity, their diameter being about 1/1000th of a millimetre (0.000039 in.); their length varied, in some cases being as much as 1/20th of a millimetre (0.0019 in.). A crowd of these long vibrios were to be seen creeping slowly along, with a sinuous movement, showing three, four, or even five flexures. The filaments that were at rest had the same aspect as these last, with the exception that they appeared punctuate, as though composed of a series of granules arranged in irregular order. No doubt these were vibrios in which vital action had ceased, exhausted specimens which we may compare with the old granular ferment of beer, whilst those in motion may be compared with young and vigorous yeast. The absence of movement in the former seems to prove that this view is correct. Both kinds showed a tendency to form clusters, the compactness of which impeded the movements of those which were in motion. Moreover, it was noticeable that the masses of these latter rested on tartrate not yet dissolved, whilst the granular clusters of the others rested directly on the glass, at the bottom of the flask, as if, having decomposed the tartrate, the only carbonaceous food at their disposal, they had then died on the spot where we captured them, from inability to escape, precisely in consequence of that state of entanglement which they combined to form, during the period of their active development. Besides these we observed vibrios of the same diameter, but of much smaller length, whirling round with great rapidity, and darting backwards and forwards; these were probably identical with the longer ones, and possessed greater freedom of movement, no doubt in consequence of their shortness. Not one of these vibrios could be found throughout the mass of the liquid.
[Illustration with caption: Figure 10.]
We may remark that as there was a somewhat putrid odour from the deposit in which the vibrios swarmed, the action must have been one of reduction, and no doubt to this fact was due the greyish coloration of the deposit. We suppose that the substances employed, however pure, always contain some trace of iron, which becomes converted into the sulphide, the black colour of which would modify the originally white deposit of insoluble tartrate and phosphate.
But what is the nature of these vibrios? We have already said that we believe that they are nothing but the ordinary vibrios of putrefaction, reduced to a state of extreme tenuity by the special conditions of nutrition involved in the fermentable medium used; in a word, we think that the fermentation in question might be called putrefaction of tartrate of lime. It would be easy enough to determine this point by growing the vibrios of such fermentation in media adapted to the production of the ordinary forms of vibrio; but this is an experiment which we have not ourselves tried.
One word more on the subject of these curious beings. In a great many of them there appears to be something like a clear spot, a kind of bead, at one of their extremities. This is an illusion arising from the fact that the extremity of these vibrios is curved, hanging downwards, thus causing a greater refraction at that particular point, and leading us to think that the diameter is greater at that extremity. We may easily undeceive ourselves if we watch the movements of the vibrio, when we will readily recognize the bend, especially as it is brought into the vertical plane passing over the rest of the filament. In this way we will see the bright spot, THE HEAD, disappear, and then reappear.
The chief inference that it concerns us to draw from the preceding facts is one which cannot admit of doubt, and which we need not insist on any further—namely that vibrios, as met with in the fermentation of neutral tartrate of lime, are able to live and multiply when entirely deprived of air.
V.—ANOTHER EXAMPLE OF LIFE WITHOUT AIR—FERMENTATION OF LACTATE OF LIME
As another example of life without air, accompanied by fermentation properly so called, we may lastly cite the fermentation of lactate of lime in a mineral medium.
In the experiment described in the last paragraph, it will be remembered that the ferment liquid and the germs employed in its impregnation came in contact with air, although only for a very brief time. Now, notwithstanding that we possess exact observations which prove that the diffusion of oxygen and nitrogen in a liquid absolutely deprived of air, so far from taking place rapidly, is, on the contrary, a very slow process indeed; yet we were anxious to guard the experiment that we are about to describe from the slightest possible trace of oxygen at the moment of impregnation.