Nov. i8. 1918 Physiological Studies of Normal and Blighted Spinach 401 



somewhat higher concentration of salts than the soil of the healthy 

 plants; other properties, however, like humus content, water-holding 

 capacity, were in favor of soil poorer in nitrogen. If any clear difference 

 existed, the soil of the diseased spinach was superior to that yielding the 

 healthy spinach. This being true, it was evident that the cause of the 

 lower nitrogen content of the diseased spinach was to be sought in the 

 plant itself. 



The loss of nitrogen may occur through denitrification, which, if it 

 does take place in the diseased spinach, would satisfactorily explain its 

 lower nitrogen content as well as other phenomena which will be men- 

 tioned subsequently in this paper. The first step in the process of 

 denitrification consists primarily in the reduction of the nitrates to 

 nitrites. The latter react on acid amids, which, as we have seen, are 

 present in the spinach tissues, whereby elementary nitrogen is set free. 

 This reaction can be presented chemically as follows : 



R.CO.NH2+NO.OH =R.CO.OH-f N2-I-H0O. 



Acid amid. Fatty acid. 



Thus, Doth the nitrogen of nitrates and of acid amids would be lost 

 through the process of denitrification, explaining the lower nitrogen 

 content in the diseased spinach. 



In the process of denitrification the reduction seems not to stop after 

 the nitrates have been reduced to nitrites, but the latter seem often in 

 part to be further reduced to ammonia. If this took place, it is evident 

 that the diseased spinach tissues would show a somewhat higher per- 

 centage of ammoniacal nitrogen than the corresponding healthy tissues. 

 This is actually the case, as a glance at the data from the winter-collected 

 material in Table VIII shows (column 7). 



Conversely, a somewhat smaller acid amid nitrogen content would be 

 expected in the diseased spinach tissues than in the corresponding 

 healthy tissues, because of the fact that the acid amids by reacting with 

 the nitrites in accordance with the supposed reaction would lose their 

 nitrogen in gas form. Evidence to support this explanation is found in 

 Table VIII (column 8). 



An examination of column 9 of the same table shows that the percent- 

 age of humin nitrogen in the diseased spinach is, as a rule, higher than 

 in the healthy spinach. Hart and Bentley {15) and Roxas (40) in Hart's 

 laboratory have demonstrated that the formation of humin nitrogen 

 takes place at the expense of diamine acids and monoamine acids. In- 

 asmuch as, under similar conditions, the humin nitrogen was formed in 

 the spinach extract by boiling with hydrochloric acid, it is evident that 

 the proportions of monoamino acids and basic nitrogen originally present 

 were higher than the values given in Table VIII (columns 10 and 11) — 

 namely, by the amount of humin nitrogen formed 

 83816°— 18 3 



