MICROBIOLOGICAL ANALYSIS OF SOIL 7l5 



NITRIFYING CAPACITY OF THE SOIL 



Several methods are available for the study of nitrification: 



1. Solution or sand method. A standard sterile solution containing a cer- 

 tain amount of ammonium sulfate and CaCOs or MgCOs as a base, in addition 

 to the necessary minerals, is placed in a series of flasks and inoculated with 10 

 per cent of the soil to be tested. The flasks are incubated at 28° to 30°C. for thirty 

 days and the nitrates formed are determined by the phenol-disulfonic acid method. 

 The results obtained by this method supply information as to the presence or 

 absence of nitrifying bacteria, influence of stimulating substances present in the 

 soil, etc. The solution method may be replaced by the sand method. One hun- 

 dred-gram portions of pure washed sand, containing 210 mgm. CaCOs are placed 

 in 250 cc. Erlenmeyer flasks; 15 cc. portions of a mineral solution (2 grams 

 K 2 HP0 4 , 1 gram MgS0 4 , 0.4 gram FeS0 4 in 1000 cc. of water) are then added to 

 each flask. The flasks are plugged and sterilized in the autoclave for 1 hour; 5 

 cc. portions of a sterile aqueous solution containing 30 mgm. of nitrogen, in the 

 form of ammonium sulfate, are added after sterilization; these are inoculated 

 with 10 grams of soil and incubated for 30 days at 28° to 30°C. The nitrates are 

 then determined. 



2. Nitrification of the soil's own nitrogen. This consists in incubating, for 

 30 days at 25° to 28°C, 100-gram portions of the soil to be tested. The soil is 

 placed in covered tumblers and contains the optimum amount of water (50 to 60 

 per cent of saturation). The results obtained by this method indicate the con- 

 dition of the forms of nitrogen present in the soil and the rapidity with which 

 these are transformed into nitrate nitrogen. 



3. Nitrification of ammonium sulfate. Thirty milligrams of nitrogen in the 

 form of ammonium sulfate are added to 100-gram portions of soil, which are 

 placed in tumblers or flasks and kept at optimum moisture for 30 days at 25° 

 to 28°C. The nitrates formed supply information on the buffer content of the 

 soil and on the maximum nitrate accumulation when no basic substances or 

 buffering agents are added. Initial and final hydrogen-ion concentrations of the 

 soil should be determined. 



4. Thirty milligrams of nitrogen as ammonium sulfate and 210 mgm. CaCOs 

 are added to the soil. The lime should be well mixed with the soil before the 

 ammonium sulfate is added. This amount of carbonate is equivalent to an 

 addition of the theoretical amount of base necessary for the complete neutraliza- 

 tion of all the nitric and sulfuric acids formed from the complete oxidation of the 

 30 mgm. of nitrogen, in the form of ammonium sulfate. The results obtained 

 by this method are more indicative of the nitrifying condition of the soil than 

 any of the other methods, since nitrification is tested here with the reaction 

 factor eliminated. Further work may, however, lead to a modification of this 

 method. Initial and final hydrogen-ion concentration should always be de- 

 termined. 



5. Nitrification of organic nitrogenous materials. One-quarter per cent of 

 organic matter with a high nitrogen content (10 to 12 per cent), such as dried 

 blood, or 0.5 to 1.0 per cent of organic materials of a low nitrogen content (cot- 

 tonseed meal, soy bean meal, alfalfa meal) should be employed. Nitrates are 



