PROTEINS 85 



crystals of impure leucine (see Fig. 145, Chapter XXV). After examining these 

 crystals under the microscope, strain off the crystalline material through fine 

 muslin, heat it gently in a little water to dissolve the leucine (the tyrosine will be 

 practically insoluble) and filter. Concentrate the filtrate and allow it to stand hi a 

 cool place over night for the crude leucine to crystallize. Filter off the crystals 

 and use them in the tests for leucine given on page 86. The crystals of tyrosine 

 remaining on the paper from the first filtration may be used hi the tests for tyro- 

 sine as given below. If desired, the tyrosine and leucine may be purified by 

 recrystallizing in the usual manner. Habermann has suggested a method of 

 separating leucine and tyrosine by means of glacial acetic acid. 



EXPERIMENTS ON TYROSINE 



Make the following tests with the tyrosine crystals prepared in 

 the above experiments, or upon those obtained during the preparation 

 of cystine (see page 86), or upon some pure tyrosine furnished by the 

 instructor. 



1. Microscopical Examination. Place a minute crystal of tyrosine on a slide, 

 add a drop of water, cover with a cover-glass, and examine microscopically. 

 Now run more water under the cover-glass and warm hi a Bunsen flame until the 

 tyrosine has dissolved. Allow the solution to cool slowly, then examine again 

 microscopically, and compare the crystals with those shown hi Fig. 25, page 75. 



2. Solubility. Try the solubility of very small amounts of tyrosine hi cold and 

 hot water, cold and hot 95 per cent alcohol, dilute NH 4 OH, dilute KOH and dilute 

 HC1. 



3. Sublimation. Place a little tyrosine in a dry test-tube, heat gently and 

 notice that the material does not sublime. How does this compare with the 

 result of Experiment 3 under Leucine? 



4. Hoffman's Reaction. This is the name given to Millon's reaction when 

 employed to detect tyrosine. Add about 3 c.c. of water and a few drops of Mil- 

 Ion's reagent to a little tyrosine in a test-tube. Upon dissolving the tyrosine by 

 heat the solution gradually darkens and may assume a dark red color. What 

 group does this test show to be present hi tyrpsine? 



5. Sulphuric Acid Test (Piria). Warm a little tyrosine on a watch glass on a 

 boiling water-bath for 20 minutes with 3-5 drops 'of cone. H 2 SO 4 . Tyrosine- 

 sulphuric acid is formed in the process. Cool the solution and wash it into a 

 small beaker with water. Now add CaCO 3 in substance slowly with stirring, 

 until the reaction of the solution is no longer acid. Filter, concentrate the 

 filtrate, and add it to a few drops (avoid an excess) of very dilute neutral ferric 

 chloride. A purple or violet color, due to the formation of the ferric salt of 

 tyrosine-sulphuric acid, is produced. This is one of the most satisfactory tests 

 for the identification of tyrosine. 



6. Formaldehyde-Sulphuric Acid Test (Morner). Add about 3 c.c. of 

 MSrner's reagent 1 to a little tyrosine in a test-tube, and gently raise the tempera- 

 ture to the boiling-point. A green color results. 



1 Morner's reagent is prepared by thoroughly mixing i volume of formalin, 45 volumes 

 of distilled water, and 55 volumes of concentrated sulphuric acid. 



