190 



albuminous matter in the nodules is so very high. 1 myself found 

 about 4V„ nitrogen, which is about 25 '/„ albumeu in the dry matter 

 of pease-nodules. Others found 5 to 6Vo nitrogen. It is noteworthy 

 that the bacterial colonies on agar plates, grown out of the plant, 

 contaiu but i to 2 "/o nitrogen of the dry weight, which consists 

 for the greater part of carbohydrates. So it is certain that the 

 bacterial body is very much modified by its entrance into the plant 

 cell as well morphologically as physiologically. Therefore it was 

 tried gazometrically to state nitrogen absorption in the tubercles. If 

 the hypothesis is founded it must be possible, with a great quantity 

 of tubercles in a closed space and under favourable physiological 

 conditions, easily to observe that absorption. For the nuniber of 

 tubercles, for example of the woody papilionaceae, being as said 

 very small, while yet these plauts are noted in agriculture for their 

 cousiderable nitrogen-Iixing power, the action of the tubercles must 

 necessarily be very intense. 



To test the hypothesis w^e acted as follows. ') P'^irst small, later 

 lai'ger quantities of lupine and serradella tubercles were placed in 

 wide glass tubes which could readily be connected with the gas 

 burettes, then put in thermostats at about 25° C. The tubercles 

 respiring vigorously we had to keep account with a rapid assimil- 

 ation and supply of the oxygen. Further it was only necessary to 

 determine the quantity of nitrogen still present after deduction of 

 the carbonic acid and the oxygen. The only difficulty we met 

 with was that the nodules, which by their abundant content 

 of albuminous matter are an excellent food for bacteria, when 

 they touch each other and get moist, easily give rise to fermentations 

 in particular by Bactermm aërogenes. Hereby hydrogen and much 

 carbonic acid are produced, so that it is then necessary also to 

 determine the hydrogen. But this fermentation may be prevented by 

 introducing the material very loosely into the burette, so that there 

 are but few points of contact between the nodules, and the air can 

 freely pass between. Under such conditions there is no danger that 

 free nitrogen will be formed ; this only occurring through the action 

 of the denitrifying bacteria on nitrates, which substance is in the 

 nodules completely absent. 



Of the tubercles of yellow lupine we used in our experiments 

 quantities of 100 grs., 500 grs., and later even of 1 kil. In some 



1) In some of these experiments I was assisted by Ir. D. C J Minkman, 

 formerly assistant to the Laboratory for Microbiology of the Technical High School 

 at Delfl. 



