142 



BUTYRIC ACID FERMENTATION 



m 



method in 1888 that it caine into general use in Bacteriology. The test-tube 

 containing the inoculated nutrient medium as a roll culture if desired is 

 placed in a large test-tube (Fig. 47) about i^ inches wide and 10 inches long, at 



the bottom of which has just 

 previously been inserted i gram 

 of dry commercial pyrogallic acid 

 and 10 c.c. of deci - normal 

 caustic potash. The smaller 

 tube rests on a small wire sup- 

 port, in order to prevent it from 

 dipping into the liquid. The 



" Buchner pyrogallol tube " is closed by a well-fitting 

 previously-moistened rubber stopper, and may then 

 be placed in the incubator. When kept at 37 C. the 

 absorption of oxygen is complete in twenty-four hours, 

 or in two days at 20 C. To treat plate cultures by 

 this method, the culture is placed over a basin contain- 

 ing a sufficient quantity of the said solution and resting 

 on a flat plate of ground glass, the whole being covered 

 with a well-sitting bell-glass, the edge of which has 

 been rubbed over with vaseline. Another method for 

 the culture of anaerobic organisms consists in expell- 

 ing the air from the culture vessel by another gas, 

 e.g. carbon dioxide, hydrogen, coal-gas, or nitrogen. 

 Carbon dioxide is frequently recommended by the 

 French school, and particularly by Pasteur, but its 

 employment is not without objections, since it is not 

 an inert gas, but is absorbed by the medium, which 

 it then renders acid, and hence has the power of 

 restricting growth. Moreover, according to the re- 

 searches of P. FKANKLAND (I.), it acts as a fatal poison 

 on many bacteria. Although experience shows that 

 hydrogen gas is not inert, still it may be accepted 

 as the best to use for anaerobic cultures. Ordinary 

 illuminating gas was recommended for this purpose 

 by R. WURTY and A. FOUREUR (I.), but, according to 

 the researches of TH. KLADAKIS (I.), it must be re- 

 jected, since he found it acting as a poison on many 

 bacteria. Nitrogen may be regarded as perfectly 

 innocuous, and would long ago have been employed 

 for anaerobic cultures were it not that the method of 

 preparation is too cumbrous and costly, at least for 

 the physiologist. Many methods have been proposed 

 for the expulsion of oxygen by one of the above 

 gases, but only two will now be briefly mentioned 

 here. That of C. FRAENKEL (IV.) is concerned with 

 the treatment of test-tube cultures, ordinary wide 

 test-tubes with two-holed stoppers being employed. 

 One of the glass tubes inserted therein reaches 

 almost to the bottom of the test-tube, whilst the second is cut off close below 

 the stopper. When filled with nutrient gelatin, agar-agar (or bouillon, 

 wort, &c.), the tubes are sterilised in the steamer in the usual way, and 

 inoculated, a current of hydrogen being introduced through the longer tube and 

 passed through. When all the air is expelled the small tubes are hermetically 



v^*^ 

 FIG. 48. 



Fraenkel's anai-robic tube. 

 Contents arranged as an Esmarch 

 culture round the walls, and 

 developed colonies appearing 

 as black spots. Somewhat 

 reduced. (After Fraenkel.) 



