282 BOTANICAL GAZETTE [OCTOBER 
The standard solutions were standardized in two ways. First, 
by titration of the alkali against standard succinic acid, which was 
prepared by dissolving 5.903 gm. of C.P. salt in 1000 gm. of distilled 
water. Second, these results were confirmed by reference to stand- 
ard hydrochloric acid whose absolute strength had been deter- 
mined by precipitation with silver nitrate and weighing of the silver 
chloride. The sodium hydroxide was found to gain strength by 
standing in glass, so this was checked every few days against the 
standard acid. 
Methyl red (45) was employed as the indicator for titrating the 
distillate. This was found to give a much sharper and more 
accurate end reaction than either cochineal or methyl orange. The 
whole of the distillate was used for titration. The results were 
found much less accurate when only part was used and computation 
made for the whole. In standardizing solutions, phenolphthalein 
was usually used as indicator, although with the strength of acids 
employed, the methyl red was found to be equally accurate. 
Culture solutions were evaporated over a water bath, down to 
about 10 cc., before adding them to the digestion flask. Previous 
to the evaporation, 5 drops of concentrated sulphuric acid were 
added to fix any free nitrogen present. This precaution applied 
especially to the cultures left exposed to laboratory air. In the 
analyses when ammonium nitrate was present, this evaporation 
was found to be especially necessary, since the heat resulting from 
the mixture of the acid and water caused a loss of nitrogen in the 
form of nitric acid or oxides of nitrogen, To avoid this, extra 
precautions were taken. After the evaporation had been carried 
to about ro cc. in the usual way, this quantity was transferred to a 
Kjeldahl flask with two or three small rinsings, and then the 
evaporation was carried nearly to dryness in this flask over a watet 
bath. It was also found necessary to have the contents of the flask 
perfectly cool before adding the mixture of sulphuric and salicylic 
acids. To accomplish this, the flask was cooled in ice water and 
then held in ice water while adding the mixture of acids. As a 
still further precaution, about 0.5 gm. of salicylic acid was added 
to the material to be analyzed and thoroughly mixed before adding 
the sulphuric-salicylic mixture. In using the ammonium nitrate 

