TOO PHYSIOLOGICAL CHEMISTRY 



If much magnesium sulphate is present a precipitate of magnesium 

 hydroxide forms which interferes with the test. If considerable 

 ammonium sulphate is present a large excess of alkali must be used. 



Testing Colored Solutions by Biuret Test. If the color of the solu- 

 tion is such as to interfere with the end-reaction of the biuret test, 

 proceed as follows: Make the solution strongly alkaline with potassium 

 hydroxide and add a solution of copper sulphate. Shake up the mixture 

 with alcohol and if protein is present the alcohol will assume the typical 

 biuret coloration. This procedure is not applicable in case the pigment 

 of the original solution is soluble in alcohol. Excess of the copper salt 

 need not be avoided in this test. 



Gies's Biuret Reagent. 1 Gies has devised a reagent for use in the biuret test. 

 This reagent consists of 10 per cent KOH solution, to which 25 c.c. of 3 per cent 

 CuS04 solution per liter has been added. This imparts a slight though distinct 

 blue color to the clear liquid. This reagent is of material assistance in performing 

 the biuret test. 



Biuret Paper of Kantor and Gies. According to Kantor and Gies 2 when 

 filter paper is immersed in the above reagent and subsequently dried it forms a 

 very satisfactory "biuret paper" which may be used in a manner analogous to 

 indicator papers. Moist papers may be used in the examination of powders which 

 are neutral or alkaline in reaction. In preparing the "biuret paper," if the filter 

 paper is left for a sufficient length of time in the reagent all traces of the copper 

 sulphate will be removed from the solution. 



5. Ring Biuret Test (Posner). This test is particularly satisfactory for use on 

 dilute protein solutions, and is carried out as follows. To some dilute egg albumin 

 in a test-tube add one-half its volume of potassium hydroxide solution. Now hold 

 the tube in an inclined position and allow some very dilute copper sulphate solution, 

 made as suggested on page 99, to flow down the side, being especially careful to 

 prevent the fluids from mixing. At the juncture of the two solutions the typical 

 end-reaction of the biuret test should appear as a colored zone (see Biuret Test, 

 page 99). 



6. The Triketohydrindene Hydrate (Ninhydrin) Reaction. To 5 c.c. of 

 dilute protein solution add one-half c.c. of a o.i per cent solution of triketo- 

 hydrindene hydrate, heat to boiling for one to two minutes and allow to cool. 

 A blue color develops if the test is positive. 



This test gives positive results with proteins, peptones, peptides, and amino 

 acids which possess a free carboxyl and a-amino group. In a concentration of 

 i per cent the ammonium salts of weak acids react positively, as do also the 

 ammonium salts of strong acids in very high concentration. Certain amines also 

 give the reaction. 3 



Proceedings of Society of Biological Chemists, Journal of Biological Chemistry, 

 7, 60, 1910. 



2 Kantor and Gies: Proc. Soc. Biol. Chem., p. n, 1910. 



3 Harding and Warnerford: Jour. Biol. Chem., 25, 319, 1916. 

 Harding and MacLean: Jour. Biol. Chem., 25, 337, 1916. 



