211 



Table 13. The presence of a lipase in the tisstie of Spongilla. ' 



In order to prove this presence I first made a sufficient (juantity 

 of pure emulsion of fat (oildrops) from the sponge tissue, in the 

 following way : 



A living Spongilla (green or colourless; or gemmulae) is rubbed and 

 pressed, the parts of the skeleton removed; then the pressed out 

 liquid is centrifuged for about 5 minutes, by which a thick mass (of 

 sponge cells and chlorophyll corpuscles) sinks to the bottom and the 

 liquid remains. The latter contains numerous oildrops (and, may be, 

 chlorophyll corpuscles). Next this liquid is evaporated at 60°; the 

 residue extracted by ether; the ether then filtered and evaporated 

 too. Then remains a substance of vaseline-like consistence, sticky, 

 with a strong smell, melting when warm and then forming a lasting- 

 greasy spot on paper, indissoluble in water, but dissoluble in ether 

 and xylol, stained red with sudan III and black with osmic acid. 

 Consequently this substance is fat. By boiling in water it becomes 

 an emulsion again, containing the same oildrop's we originally pro- 

 ceeded from. (Besides, this boiling is necessary to destroy all traces of 

 enzymes, that miglit still ])e present.) I shall call this boiled liquid 

 ,, emulsion". 



Next anotlier living sponge is rubbed and pressed, etc., etc. (see 

 above); while the liquid, remaining after centrifuging, is kept. This 

 will contain the lipase, at least when it is present in sponge tissue. 

 This liquid I shall call ,, enzyme". 



As one knows, the lipase splits the fat by hydrolysis into its com- 

 ponent parts: the glycerine and the acids. It is the arising of the 

 latter we have to show in our experiments; in the following way: 

 A certain quantity of ,, enzyme" and ,, emulsion" are mixed; to this 

 we add one drop of the indicator phenolphthaleine — being red in 

 alkaline milieu, but colourless in an acid one — and such a small 

 quantity of an (alkaline) Na^ COj solution that the whole mixture 

 becomes light-red. Tlie acids, then, set free from the ,, emulsion" by 

 the lipase will make the red colour disappear. In order to get a pure 

 result it is necessary, however, that in this mixture no acids arise in 

 another way than by the hydrolysis due to the lipase; or at least 

 that we reckon with it, if it proves to be the case. 



3 Series of experiments (I, II and III) were made at the same time, 

 in which the following substances were mixed: 



[. 15 drops of ,, enzyme" -\- 2.5 cM^ of ,, emulsion" -|- 1 drop of phenolph. -f- Na^ CO3 sol. 

 [. 15 drops of ,, enzyme" -\- 2.5 cM' of water -f- 1 drop of phenolph. -f- Na^ CO3 sol. 



I. 15 dro{)s of water -f" 2.5 cM'' of ,, emulsion" -|- 1 drop of phenolph. -|- Naj CO3 sol. 



