68 THE GENERAL CHARACTERS OF THE PROTEINS 



nitroso bodies are formed, which on hydrolysis might be expected to 

 yield the same products as the substances from which they are 

 formed, with evolution of ammonia (Paal). 



On the other hand, the free amino groups in the molecule, as well 

 as the amide-nitrogen groups, might be attacked, as in the case of 

 asparagine 



2 C 4 H 8 N 2 3 + 2 HN0 2 = 2C 4 H 6 5 + 2 N 2 + 4 H 2 O. 



The amount of nitrogen evolved might serve as a measure of the free 

 amino and amide groups in a protein. It must be remembered, 

 however, that the polypeptides also act on nitrous acid in a some- 

 what indefinite manner (Fischer and Koelker) ; furthermore, in highly 

 complex substances, such as the proteins, it is also conceivable that 

 only a limited number of amino or amide groups may be the subject 

 of attack, owing to stereo-chemical reasons. Nevertheless, the amount 

 of nitrogen eliminated by nitrous acid treatment under certain speci- 

 fied conditions may be a fixed quantity and a characteristic of each 

 individual protein, and the determination of this so-called amino- 

 index has formed the subject of recent investigation by Horace 

 Brown and his co-workers. It has so far only been applied to a 

 limited number of proteins. When a pure amino-acid, such as 

 aspartic acid, is treated with nitrous acid, twice as much nitrogen is 

 evolved as the acid itself contains, one-half being derived from the 

 nitrous acid, e.g. : 



C 4 H 7 O 4 N + HNO 2 = C 4 H 6 O 5 + N 2 + H-jO. 1 



In the case of proteins, therefore, one-half the nitrogen evolved, 

 expressed as a percentage of the total nitrogen, gives the apparent 

 proportion of the nitrogen present in the amino form, or, at any rate, 

 that proportion which will react with nitrous acid. This is termed by 

 Brown the amino-index, and is represented by the symbol Aol. 

 The following are the amino numbers for a series of proteoses and 

 peptones obtained by fractionating malt proteoses : 



Malt proteoses I. 4/0 



II. 5-o 



III. 20'0 



Malt peptone I. 10*9 



II. 19-3 



Method. 



The method of determining the amino-index adopted by Brown 

 and his co-workers is a modification of that previously employed by 

 Sachsse and Kormann for determining the amino-acids present in 

 certain technical samples. The principle consists in the treatment 

 of the substance under examination with nitrous acid in statu nascendi, 

 and the measurement of the nitrogen evolved after absorption of the 

 surplus nitric oxide carried over by the gas. Certain errors were 

 found by Brown to be inherent in the method as originally suggested 

 by Sachsse and Kormann. The chief sources are due to (i) the 

 residual air in the apparatus, or dissolved in the liquid ; (2) difficulties 

 attending the production of carbonic acid of high degree of purity 

 when this is employed for freeing the apparatus from air ; (3) diffi- 

 culties associated with the complete absorption of nitric oxide with 



1 With asparagine containing an amido group only relatively half as much nitro- 

 gen is evolved. See equation already given above. 



