156 PHYSIOLOGY OF NUTRITION 



amounts of proteins and these are the regions of most active growth. The 

 unshaded parts represent fully grown tissues, which contain only a very small 

 amount of proteins, these substances, having disappeared during the growth 

 process. An exception to this statement are full-grown leaves, which contain 

 much protein material in their chloroplasts. These diagrams show not only 

 the protein contents but also the growth activities of the different parts of the 

 plant. 



The principal chemical reactions of proteins are given below. a 



1. With copper sulphate and caustic potash solution, a dark violet color is 

 produced (biuret reaction). Albumoses and peptones give a red color with this 

 reagent. This reaction is of special importance, since it serves as a means of 

 distinguishing the albumins from their cleavage products. Excess of copper 

 sulphate is to be avoided, since the blue color of this salt may obscure the 

 result. 



2. Heating with strong nitric acid gives a deep yellow color, which changes 

 to orange-red upon treatment with an excess of ammonia (xanthoproteic reac- 

 tion). 



3. Heating with Millon's reagent gives a red color (Millon's reaction). 



4. With a-napthol and concentrated sulphuric acid a blue-violet color is 

 produced (Molisch's furfurol reaction). 



5. Boiling with fuming hydrochloric acid gives a bluish-violet color (Lieber- 

 mann's reaction). 



The following additions may be useful. (1) For the biuret test, add strong KOH solu- 

 tion and follow with weak CuS04 solution. A partially decomposed protein — such as pep- 

 tones — gives a pink or purplish-red color. Gies and Kantor give directions for a single solution 

 suitable for this test. (Gies, W. J., and Kantor, J. L., Methods of applying the biuret test. 

 Biochem. bull 1: 264-269. 1911.) To 1000 cc. of 10-per cent, aqueous solution of NaOH 

 add 25 cc. of a 3-per cent, solution of CuSO*, a few cubic centimeters at a time, with thorough 

 shaking after each addition. Filter through glass wool if necessary. The biuret test, as 

 well as many other microchemical reactions, may be influenced by other reactions, the possible 

 occurrence of which must be considered. (See: Mathewson, C. A., A study of some of the 

 more important biochemical tests. Biochem. bull. 2: 181. 1912.) Biuret is represented 

 by the formula, NH 2 CO-NH-CONH 2 . In treating sections, a solution of copper hydrate 

 in KOH solution may be used. It is sometimes better to warm the section in weak KOH 

 solution, wash in water and treat with CUSO4 solution, after which it is again washed and 

 then examined in KOH solution. (3) Millon's reagent is a solution of mercuric nitrate and 

 nitrous acid. To prepare it, dissolve (in fume cupboard) mercury in twice its weight of strong 

 HNO3 (spec. grav. 1.42), and then dilute the solution to three times its volume, with water. 

 This reaction and the xanthoproteic reaction are dependent on the tyrosin or tryptophan group 

 in the protein molecule. (4) For the furfurol reaction, add a few drops of 10-20-per cent, alco- 

 holic solution of et-naphthol, and then slowly add concentrated H2SO4. The color reaction 

 appears at junction of the two liquids. If thymol is used instead of a-naphthol a carmine color 

 is produced. (5) Liebermann's reaction is used with material that has previously been 

 extracted with alcohol and ether, and it appears to be due to glyoxylic (glyoxalic) acid present 

 in the ether, this acid reacting with the tryptophan group of protein. (6) For Adamkiewicz's 

 reaction, material is extracted with ether to remove fat, dried and then extracted with glacial 

 acetic acid. The concentrated H2SO4 is added slowly and the color appears at the junction of 

 the two liquids. Here, also, the reaction seems due to glyoxylic acid (present in the acetic 

 acid). — Ed. 



