114 



A SYMPOSIUM ON RESPIRATORY ENZYMES 



nucleosidase, but contain some nuclease, which causes splitting into 

 the mononucleotides, and a nucleotidase, which removes the phos- 

 phoric acid. Table 5 illustrates the course of the enzymatic hydrolysis 

 of cozymase. 



The isolation of the nicotinamide nucleoside is complicated by 

 the fact that thus far only one reagent has been found which pre- 

 cipitates the compound, namely, phosphotungstic acid. The isolation 

 and purification, therefore, as given in Table 6, consists mainly in 

 removing the other compounds and impurities from the mixture. 



The nicotinamide nucleoside gives no characteristic precipitates 



Table 6.— Preparation of nicotinamide nucleoside 



COZYMASE 



enzymatic hydrolysis at pH 4.5 



(Nicotinamide nucleoside 

 UYDROLYS ATE\ Adenosine 



I Phosphoric acid 

 [Protein 

 dialysis; Ba(OH)2(P04i) 



SOLUTION (nicotinamide nucleoside, adenosine) 



+ Ag2S04 



SOLUTION (nicotinamide nucleoside) 

 +H2S (Ag2S i ) 

 -f-phosphotungstic acid 



PRECIPITATE SOLUTION 



suspended (discarded) 



in dil. H2SO4, 



phosphotungstic acid removed 

 by amyl alcohol-ether 

 extraction 



SOLUTION 



+picric acid (impurities [ ) 



ether extraction; 



precipitation by acetone -|- ether 



NICOTINAMIDE NUCLEOSIDE (crude product) 

 filtration through 

 aluminum oxide; 

 fractional precipitation 



NICOTINAMIDE NUCLEOSIDE 



PRECIPITATE (adenosine) 



-f-HsS (Ag2S i ) 

 -|-picric acid 



ADENOSINE PICRATE 



ether extraction; 



repeated 



recrystallization 



ADENOSINE 



