BILE 



209 



(d) Surface Tension Test (Hay). This test is based upon the principle that 

 bile acids have the property of reducing the surface tension of fluids in which they 

 are contained. The test is performed as follows : Cool about 10 c.c. of diluted 

 bile hi a test-tube to i7C. or lower and sprinkle a little finely pulverized sulphur 

 upon the surface of the fluid. The presence of bile acids is indicated if the 

 sulphur suiks to the bottom of the liquid, the rapidity with which the sulphur sinks 

 depending upon the quantity of bile acids present in the mixture. The test is said 

 to react with bile acids when they are present in the ratio of i : 120,000. 



(e) Neukomm's Modification of Pettenkofer's Test. To a few drops of 

 diluted bile in an evaporating dish add a trace of a dilute sucrose solution and one 

 or more drops of dilute sulphuric acid. Evaporate on a water-bath and note the 

 development of a violet color at the edge of the evaporating mixture. Discontinue 

 the evaporation as soon as the color is observed. 



(f) Peptone Test (Oliver). To 5 c.c. of diluted bile add 2-3 drops of acetic 

 acid, filtering if necessary. Add an equal volume of a i per cent solution of 

 Witte's peptone to the acid solution. A precipitate is produced which is insoluble 

 in excess of acetic acid. This precipitate is a compound of protein and bile acids. 



6. Crystallization of Bile Salts. To 25 c.c. of undiluted bile in an evaporating 

 dish add enough annual charcoal to form a paste and evaporate to dryness on a 

 water-bath. Remove the residue, grind it in a mortar, and transfer it to a small 

 flask. Add about 50 c.c. of 95 per cent alcohol and boil on a water-bath for 20 

 minutes. Filter, and add ether to the filtrate until there is a slight permanent 

 cloudiness. Cover the vessel and stand it away until crystallization is complete. 

 Examine the crystals under the microscope and compare them with those shown in 

 Fig- 55> page 205. Try one of the tests for bile acids upon some of the crystals. 



7. Analysis of Biliary Calculi. Grind the calculus in a mortar with 10 c.c. 

 of ether. Filter. 



Filtrate I. 



Add an equal volume of 95 per cent alco- 

 hol 1 to the ether extract, allow the mix- 

 ture to evaporate and examine for choles- 

 terol crystals (Fig. 57, page 210). (For 

 further tests see Experiment 8, p. 210.) 



lue I. 



(On paper and in mortar.) 



Treat with dilute 

 filter. 



lydrochloric acid and 



Filtrate II. 



Test for calcium, phosphates, and iron. 

 Evaporate remainder of filtrate to dry- 

 ness in porcelain crucible and ignite. 

 Dissolve residue in dilute hydrochloric 

 acid and make alkaline with ammonium 

 hydroxide. Blue color indicates copper. 



Residue IL 



(On paper and in mortar.) 

 Wash with a little water. Dry the filter paper. 



Treat with 5 c.c. chloroform and filter. 

 I 



Treat with 

 alcohol. 



5 c.c. 



of hot 



Biliverdin. 



1 The alcohol is added because of the fact that it is often found that crystallization from 

 pure ether does not yield typical cholesterol crystals. 



