ENZYMES AND THEIR ACTION 13 



5. Demonstration of a Vegetable Protease. A commercial preparation of 

 papain (papayolin, carase or papase), the protease of the fruit of the pawpaw (carica 

 papaya), may be used in this connection. Follow the same procedure as that de- 

 scribed under Gastric Protease (see p. 12). 



It has been demonstrated by Mendel and Blood 1 that the presence of HCN 

 will accelerate the proteolytic activity of papain. It is suggested that the HCN 

 acts as a so-called co-enzyme (see page 7). 



Vines 2 believes that "papain" consists of a mixture of two enzymes, a pepsin 

 and an erepsin. Mendel and Blood do not consider the evidence on this point as 

 conclusive. 



m. LIPASES 



1. Preparation of Pancreatic Lipase. 3 An extract of this enzyme may be 

 prepared from the pancreas of the pig or sheep according to the directions given 

 on page 192.* 



2. Demonstration of Pancreatic Lipase. Into each of two test-tubes intro- 

 duce 10 c.c. of milk and a small amount of litmus powder. To the contents of one 

 tube add 3 c.c. of a neutral extract of pancreatic lipase and to the contents of the 

 other tube add 3 c.c. of a boiled neutral extract of pancreatic lipase. Keep the 

 tubes at 38C. and watch for color changes. 



The blue color of the litmus powder will gradually give place to a 

 red. This change in color of the litmus from blue to red has been 

 brought about by the fatty acid which has been produced through the 

 lipolytic action exercised by the lipase upon the milk fats. 



3. Preparation of Vegetable Lipase. This enzyme may be readily prepared 

 from castor beans, several months' old, by the following procedure. 6 Grind the 

 shelled beans very fine 6 and extract for twenty-four-hour periods with alcohol-ether 

 and ether, in turn. Reduce the semi-fat-free material to the finest possible consist- 

 ency by means of mortar and pestle and pass this material through a sieve of very 

 fine mesh. Place this material in a Soxhlet extractor and extract for one week. 

 This fat-free powder may then be used to demonstrate the action of vegetable 

 lipase. Powder prepared as described may be used in quantitative tests. For 

 ordinary qualitative tests it is not. necessary to remove the last traces of fat and 

 therefore the extraction period in the Soxhlet apparatus may be much shortened. 



4. Demonstration of Vegetable Lipase. The lipolytic action of the lipase pre- 

 pared from the castor bean, as just described, may be demonstrated in a manner 

 entirely analogous to that used hi the Demonstration of Pancreatic Lipase, see 

 above. Proceed as indicated in that experiment and substitute the vegetable 

 lipase powder for the neutral extract of pancreatic tipase. The type of action is 

 entirely analogous in the two instances. 



1 Mendel and Blood: Journal of Biological Chemistry, 8, 177, 1910. 



2 Vines: Annals of Botany, 19, 174, 1905. 



'Also called steapsin. For a discussion of the enzyme see p. 191. A very active lipo- 

 lytic extract may also be prepared from the liver. 



4 If preferred, a glycerol extract may be prepared according to the directions given by 

 Kanitz (Zeitschrift fur physiologische Chemie, 46, 482, 1906) or commercial pancreatin 

 may be employed. 



5 A. E. Taylor: On Fermentation; University of California Publications, 1907. 



6 The shells should be removed without the use of water. These beans are poisonous, 

 due to their content of ricin. 



