440 PROTEINS 



adding concentrated sulphuric acid so that the latter forms a 

 separate layer at the bottom of the test tube. After a short 

 time a pink ring is produced at the junction of the two liquids, 

 and on careful agitation the colour extends over the whole 

 solution. 



According to an experiment devised by Molisch, the above 

 tests may be employed for demonstrating the distribution of 

 protein in leaf tissue. The leaf is freed from chlorophyll by 

 steeping in boiling water and then in boiling 75 per cent 

 alcohol ; separate portions of the leaf are then left to soak for 

 half an hour in [a] nitric acid (i : 2), {b) Millon's reagent, and 

 [c) 5 per cent copper sulphate. Portion (a) is then transferred 

 to a solution of ammonia, while portion [c) is placed in 10 

 per cent caustic potash. The characteristic colour changes 

 are produced. 



Microchemical Reactions. 



The following are the usual microchemical tests employed 

 for the indication of proteins within the plant : — 



1. Iodine gives a yellow to brown coloration. 



2. With osmic acid a brown coloration results. 



3. Biuret Reaction. — A solution of copper hydrate in caustic 

 potash may be added direct to the preparation ; or the section 

 may be steeped for some time, say twenty to sixty minutes, in 

 0-2 per cent solution of potash, washed, placed in a 10 per cent 

 solution of copper sulphate for thirty to sixty minutes, washed 

 in water and mounted in a 2 per cent solution of caustic 

 potash. A mauve to violet coloration indicates the presence 

 of proteins. 



4. Millons Reagent. — The section or scraping is mounted 

 in a few drops of the reagent and warmed. A brick-red 

 coloration results when proteins are present. 



5. Xanthoproteic Reaction. — A yellow to orange colora- 

 tion results with proteins. The preparation is warmed on the 

 slip with a few drops of strong nitric acid. The proteins 

 acquire a yellow colour which is changed to orange on 

 moistening with strong ammonia. 



