INULASE FORMATION IN ASPERGILLUS 85 



Various methods for inoculation of the cultures with fungus, 

 spores were tried out with the \'iew of obtaining the most uni- 

 form growth possible of the fungus and at the same time avoiding 

 chances of contamination of the cultures. The beet results from 

 all points of view were obtained by transferring two loopfuls of 

 spores to each culture flask with a standard platinum loop. The 

 flasks were then thoroughly agitated to give a uniform distri- 

 bution of the spores. 



All cultures were grown in the dark in a room regulated by an 

 electrical thermostat at a temperature ranging from 29.5° to 

 30°C. Under such conditions, Aspergillus niger on a 1% inulin 

 mediun begins to form mature spores after not more than forty- 

 eight hours. At the end of thirty-two hours conidiophores be- 

 gin to form but cultures are still white and no mature spores are 

 present. On media containing other sources of carbon than 

 inulin, Aspergillus niger takes somewhat longer to sporulate. 

 Other fungi grown on the same media and under the same condi- 

 tions vary considerably in the rapidity of their growth and 

 maturation. 



As will be seen from the review of the literature and from the 

 experimental data which will be considered later, the period at 

 which the mycelium is richest in enzyme content, is at the be- 

 ginning of sporulation. This is apparently the time of greatest 

 metabolic activity of the fungus. At this time the mycelium was 

 removed and washed until tests with Fehling's solution no longer 

 showed the presence of reducing sugar from the culture medium, 

 in the wash water. It is believed that this amount of washing 

 was sufficient to remove all but traces of the culture medium 

 from the mycelium. The mycelium was then pressed between 

 pieces of filter paper to remove the free water present and treated 

 according to a modification of the method used by Bartholomew 

 (1914) in his study of the enzymes of certain marine algae. 

 This method consists in immersing the water free mycelium for 

 twenty minutes in 95% alcohol, then removing the excess of 

 alcohol by means of filter paper and immersing it in acetone for 

 ten minutes and a second time for two minutes. The excess of 

 acetone having been removed, the material was spread out and 



