274 PHYSIOLOGY OF THE FUNGI 



ENZYME PRODUCTION 



The industrial production and use of enzymes from microorganisms 

 in the Occident is fairly recent, although the use of fungi as amylolytic 

 agents by the peoples of the Orient for the preparation of koji and other 

 foods is an old art. For this purpose, mixed cultures of species of Asper- 

 gillus and Rhizopus are grown upon the rice or soybean substrates, the 

 enzymes being used without separation. The pioneering work of Taka- 

 mine (1914) on the amylases of A. oryzae was especially important. 



The ability of fungi to produce amylase is widely distributed, but only 

 a few species are used commercially for this purpose. The amount of 

 amylase produced varies with the species or isolate and the cultural condi- 

 tions. Le Mense et al. (1947) screened 359 isolates of Penicillium and 

 Aspergillus and found 42 isolates to produce amylase in submerged 

 culture. The activity of the species of Penicillium ranged from 0.1 to 

 0.6 enzyme unit per milliliter of culture medium. One isolate of A. 

 niger (NRRL 337) was found to be especially adapted for the production 

 of amylase in submerged culture. The production of amylase was highly 

 dependent upon the composition of the medium. Corn meal was espe- 

 cially valuable in increasing amylase production when added to basal 

 media composed of corn steep liquor, dried tankage, soybean meal, or 

 thin stillage. Amylase production was stimulated by the addition of 10 

 to 20 mg. of sodium chloride per liter of culture medium. Addition of a 

 mixture of chlorinated phenols (Dowcide G) inhibited sporulation and 

 increased amylase production (Erb et al., 1948). 



Others have found different isolates of the same species to produce 

 varying amounts of amylase. Hao et al. (1943) studied the production 

 of amylase by 27 isolates of various species of fungi when grown upon 

 wheat bran. A. oryzae, Rhizopus delemar, and R. oryzae produced the 

 largest amounts of amylase. A. oryzae was the fungus of choice because 

 of ease of handling. 



In practice, fungus amylases are produced and utilized in three general 

 ways. (1) In the amylo process, starch is solubilized by autoclaving with 

 a trace of a mineral acid, and the mash is inoculated with a species of 

 Rhizopus, which produces amylase abundantly, and a species of yeast. 

 The function of the Rhizopus species is to convert the starch into ferment- 

 able sugars, from which the yeast produces alcohol. For a description 

 of this process see Owen (1933). (2) The fungus may be grown upon a 

 solid substrate such as bran and the resulting moldy mass (mold bran) 

 dried (Underkofler et al., 1946). The fresh material may be used without 

 drying (Roberts et al., 1944). (3) Fungus amylases may be produced in 

 submerged aerated cultures much as antibiotics are produced. The 

 culture medium may be used directly to replace malt as a saccharifying 

 agent. 



