GAS PRODUCTION 289 



4. Note spectroscopic reactions. 



7. Reducing Agent Formation. 



(a) Colour Destruction. 



1 . Prepare tube cultivations in nutrient bouillon tin- 

 ted with litmus, rosolic acid, neutral red, and incubate. 



2. Examine the cultures each day and note whether 

 any colour change occurs. 



(b) Nitrates to Nitrites. 



Medium Required: 



Nitrate bouillon (vide page 185). 



Or nitrate peptone solution (vide page 186). 

 Reagents Required: 



Sulphuric acid (25 per cent.). 



Metaphenylene diamine, 5 per cent, aqueous solution. 



METHOD. 



1. Prepare tube cultivations and incubate together 

 with control tubes (i. e., uninoculated tubes of the 

 same medium, placed under identical conditions as to 

 environment) . 



This precaution is necessary as the medium is liable 

 to take up nitrites from the atmosphere, and an 

 orjfinion as to the absence of nitrites in the cultivation 

 is often based upon an equal colouration of the medium 

 in the control tube. 



Test both the culture tube and the control tube for 

 the presence of nitrites. 



2. Add a few drops of sulphuric acid to the medium 

 in each of the tubes. 



3. Then run in 2 or 3 c.c. metaphenylene diamine 

 into each tube. 



Brownish-red colour = nitrites. 

 The depth of colour is proportionate to the amount 

 present. 



8. Gas Production. 



(A) Carbon Dioxide and Hydrogen. 

 Apparatus Required: 



Fermentation tubes (vide page 161) containing sugar bouillon 



