100 tapwater 
0,5 fat *) (finely divided) 
0,5 caleiumearbonate 
0,05 kaliumsulphate 
0,1 magnesiumammonium-phosphate. 
In this feebly alkaline medium the fat-splitting bacteria grow very 
well; the chalk and the magnesiumammonium-phosphate serve to 
neutralise the fatty acid and the acids resulting from glycerin. 
Aërobie culture at 18°—25° C. If we inoculate a layer of the 
above medium, + one centimeter thick, in an ERrLENMEYER flask, 
with garden soil, sewage mud, canalwater or dung, and cultivate 
at 18°—25° C., a rapid increase of the bacteria introduced with 
the inoculation material ensues. After one or two transferences to 
a same medium there is an abundance of these microbes. The 
changes observed in the medium are the following: After one or 
two days the liquid becomes turbid by the bacterial growth and 
usually assumes a yellowish green colour; the floating pieces of fat 
sink down on saponification and subsequently change into slimy 
flakes. 
Most of the bacteria present in the medium belong to the fat- 
splitting species; among them are melting and non-melting micro- 
cocci and fluorescents and species corresponding to B. punctatum. 
As well among the fluorescents as among the last named, stocks 
are found that split fat very vigorously, whereas others do this 
feebly or not at all. 
If we take fatty acid instead of fat for source of carbon, the 
same flora occurs, whilst with glycerin very fine accumulations of 
jluorescents are obtained. 
Avrobic culture at 30°—87° C. At these temperatures the culture 
presents quite another aspect as at those between 18°—25°. Thus 
we often find on the liquid a film of Sperillum especially when inocula- 
ting with sewage or canal water; it gets, however, lost after one or 
two transplantations. Evidently the spirilla have no lipolytic enzymes 
and grow at the expense of the products formed by other microbes. 
Sometimes a not inconsiderable growth of bay bacteria and butyric 
1) The fat used for these experiments is the so-called “suif pressé”, a product 
remaining behind after pressing the oleo margarine from tallow. The melting- 
point is + 55°C., the saponificationnumber 193—195. In consequence of the high 
melting point it is easy finely to distribute this fat in the culture liquid by shaking it 
in melted state with the latter and quickly cooling it. Also for culture at high 
temperatures (+ 52°), for anaérobic culture and for experiments on denitrification 
with fats, it shows advantages over easily melting fats, 
