I04 Journal of Agricultural Research voi. xv, No. a 



part in a warm compartment of the greenhouse, part in a cold frame 

 at temperatures usually above o° C, but frequently low enough to 

 freeze the plants stiff. The hardened plants were uninjured by an 

 hour's freezing at —3°, while those from indoors were killed. Samples 

 of juices were collected from the leaves of both hardened and non- 

 hardened plants, using the same method for grinding and expressing 

 in both cases. The juices were quickly frozen in Nessler tubes in a 

 freezing mixture at —4° and kept at that temperature for two hours. 

 Control samples were placed at once into ice water and kept there for 

 the same time as the frozen samples. After two hours the frozen samples 

 were quickly thawed in slightly warm water and then cooled to 0° as 

 soon as all the ice had melted. All the samples were then placed in 

 tubes and centrifuged together for eight minutes at high speed. Sam- 

 ples were withdrawn with a pipette to avoid disturbing the precipitates. 

 The total nitrogen was determined by the Nitrogen Laboratory of the 

 Bureau of Chemistry. On expressing the total nitrogen as proteins, it 

 was found that 9.4 per cent of the original quantity of protein was 

 precipitated from the hardened plants, and 31.2 per cent from the non- 

 hardened plants by this period of freezing. 



A change in state of proteins sufficient to allow these proportions to 

 be precipitated by centrifuging the juices certainly bears some relation 

 to frost injury. The lesser amount of precipitation in the case of juices 

 from hardened cabbages indicates that this may account for the lesser 

 injury to such plants. 



PRECIPITATION OF THE PROTEINS OF HARDENED AND NONHARDENED 

 CABBAGES ON THE ADDITION OF ACID 



Juices were obtained in the same manner as before from hardened and 

 nonhardened cabbages. To 50 cc. samples a sufficient quantity of acid 

 was added to increase the hydrogen-ion concentration by approximately 

 the amount by which it was increased by freezing at —3° C. This 

 quantity was found by previous expeiiments to be 5 cc. of N/io sul- 

 phuric acid for 50 cc. of juice from nonhardened plants. The juices 

 were kept in ice water for an hour, together with controls. The sam- 

 ples were then treated as in the previous experiment. It was found that 

 this quantity of acid precipitated 11 per cent of the original quantity 

 of proteins in the juice from hardened plants and 44 per cent in the 

 juice from nonhardened plants. This indicates that the proteins are 

 more easily precipitated by increase of the hydrogen-ion concentration 

 in the juices of nonhardened plants than of hardened plants. This sen- 

 sitiveness of the proteins to the addition of acid is closely paralleled by 

 their sensitiveness to freezing. 



