2<5o PRINCIPLES OF GENERAL PHYSIOLOGY 



beer wort, the growth is vigorous. The effect of adding larger quantities is said 

 to be due to the presence of a substance which Wildiers calls " bios," provisionally, 

 until its chemical nature is known. This bios is found in all cultures of growing 

 yeast. If the culture medium is inoculated with an insufficient quantity of 

 yeast, and, at the same time, with small quantities of boiled yeast extract, even 

 if filtered through porous clay, growth is ensured. The beneficial effect of 

 phosphate on fermentation by yeast is well known, so that it is necessary to note 

 that the artificial culture fluid used contained this salt already. A culture 

 medium may sometimes become infected with yeast from the air, and it seems 

 difficult to understand how bios could be taken with it ; however, according to 

 the observations of Kossovics (1896), it seems possible that bacteria might produce 

 some substance of this nature. Since this work seems to have been somewhat 

 overlooked, it is advisable to give some further details. It is interesting to note 

 that the work was done at the University of Louvain, whose tragic fate has 

 aroused the indignation of the whole civilised world. 



As to the chemical nature of bios, we may note that it is soluble in 80 per cent, alcohol, not 

 in absolute alcohol nor in ether. It is not precipitated by lead acetate, phospho-molybdic nor 

 phospho-tungstic acid, nor by mercuric chloride. It is present in Liebig's meat extract and 

 commercial peptone as well as in decoction of germinated malt (beer wort) before the action of 

 yeast. It is not contained in the products of peptic or tryptic digestion of pure proteins. 

 Thymus nucleic acid does not give the result. Whatever it may be, it is evidently not the 

 same substance as that of Hopkins, since it is insoluble in ether, nor is it Funk's " vitamine " 

 since it is not thrown down by precipitants of organic bases. An important point is that it is 

 not produced by the yeast plant itself in the course of its growth, in fact it seems to disappear. 

 If the culture to which originally a certain amount of bios was added be boiled and 

 concentrated, it is found necessary to add at least that amount of the concentrated extract 

 which would contain the original quantity of bios in order to ensure growth in a new 

 experiment. Wildiers calls attention to the circumstance that the results seem to give 

 a possible explanation of the famous contest between Liebig and Pasteur. The latter 

 inoculated his cultures with a fragment of yeast of the size of the head of a large pin. This 

 was probably about the lowest limit of "bios," so that, if Liebig took a smaller amount, he 

 would not get growth. As Wildiers remarks (p. 328), if Liebig had accepted Pasteur's 

 invitation to see his experiments, the discrepancy would probably have been explained. We 

 see also how minute is the amount of bios required. A smaller quantity than the optimal 

 will allow of a very slow growth, but, under the microscope, there are to be seen, at one time, 

 very few living cells, and it appears that, in order that a new cell may grow, it has to wait for 

 the death of an old one in order to obtain the bios required. On the whole there appears to 

 be some essential structural unit which the yeast cell is unable to form for itself. 



Abel Amand (1903) answers some possible objections to these experiments. We saw that 

 the influence of the number of cells was eliminated in that the same number was ineffective in 

 Pasteur's medium, effective in malt extract. Amand 's experiments were made to test the 

 suggestion that the bios added was effective by counteracting some poisonous substance in the 

 water used for the culture fluid, some " oligodynamic action" as described above (page 222). 

 Copper was suggested. The water from glass stills, however, gave the same results. The 

 chemicals used were also tested, and, although the details must be read in the original paper, 

 the possibility of poisonous constituents seems satisfactorily excluded. Moreover, one may 

 recall the experiments of Ringer, in which the poisonous action of distilled water was 

 abolished by calcium, which is a constituent of the yeast culture medium. Amand, in a 

 further paper (1904), shows that bios disappears and cannot be extracted from the cells, 

 so that it is either used for synthesis of a more complex substance or undergoes spontaneous 

 decomposition. According to Devloo (1906) the active principle is a base, precipitated by 

 mercuric chloride in presence of barium hydroxide. It can be made to replace choline, at all 

 events partially, in lecithin. It may naturally occur in the form of the base of a lecithin-like 

 substance. It is not choline itself nor can it be prepared from choline. 



Twort and Ingrain (1912) have made the interesting observation that Johne's 

 bactthis, responsible for the pseudo-tubercular enteritis of oxen, will "only grow 

 on a medium to which dead tubercle bacilli, or some other " acid-fast " bacilli, 

 have been added. These authors have also been able to extract by alcohol 

 this essential substance. 



The work of Bottomley (1914) is also of interest. He finds that substances 

 which stimulate plant growth are formed in sphagnum peat when it is incubated 

 with cultures of certain aerobic soil" bacteria. They are very active ; the watery 

 extract of 0'18 g. of peat thus treated caused the growth of Primula seedlings 

 in six weeks to be double that of control plants. They seem to be allied to 

 the " accessory factors " of diet. 



Thornton and Smith (1914) find that the chlorophyll-containing protozoon, 



