45 
Exercise 23. Hydrogen Sulphide Formation from 
Protein 
MATERIALS: 
3 tubes of egg gelatin 
3 tubes of egg agar 
Garden soil 
Cultures of: Escherichia coli 
Bacillus mycoids 
Bacillus grave ole ns 
1. Observe the precautions in handling gelatin, and prepare a stab culture 
of each of Escherichia coli , Bacillus mycoides and Bacillus gravcolens. 
2. Incubate the cultures at 20° C. 
3. Melt the agar and add about 0.1 gram of sterile lead carbonate to each 
tube. Mix well by rolling the tubes between the hands. 
4. Inoculate one tube, after cooling to 45° C. with a loopful of garden soil. 
Mix well. Inoculate the second tube with one loopful from the first. Mix 
well. Inoculate the third tube from the second, using three loopfuls. 
5. When the agar is solid incubate the culture at 30° C. 
6. After one week examine the agar shake cultures for black colonies, and 
the gelatin stab cultures for liquefaction. 
7. If the gelatin has liquefied, notice the odor and add a few drops of 5 per 
cent lead acetate solution to each tube. Observe the results? 
(a) What is the black substance formed upon the addition of 
lead salts? 
( b ) What is the source of the hydrogen sulphide? 
(c) How do you explain the power of these organisms to liquefy 
gelatin and produce “lead blackening”? 
( d) From what compound in the media is the H s S produced? 
(e) Where do these organisms occur in nature? 
(0 What is the practical value of these organisms? 
