Production of industrial enzymes in solid state fermentation of agricultural wastes by zygomycetes

Abstract

Large amount of corn-stalks and leaves arise as agricultural waste during the agro-industrial
processes. Solid-state fermentation to produce different industrial enzymes such as cellulolytic
enzymes, lipases and proteases, and bioconversion to ethanol may provide an alternative and
economic path to utilize these residues. Zygomycetes fungi have been assumed to play an
important role in the decomposition of plant and other organic materials in consequence of
their effective extracellular enzyme production. Several members of them are commonly used
in different biotechnological applications for the large scale production of industrial enzymes,
however, production of these enzymes on agricultural wastes are poorly characterized until to
date. The aim of our present study was to investigate the activities of cellulolytic enzymes and
the production of lipase and protease by zygomycetous strains on agro-industrial wastes such
as corn-stalks, corn-leaves and wheat bran.
The Mucor, Rhizomucor, Gilbertella and Rhizopus isolates selected for this study have proven
to be good extracellular β-glucosidase producer in our previous experiments in which
submerged cultivation on cellobiose and solid-state fermentation on wheat bran were used
(TAKÓ et al, 2010). In these studies, solid-state fermentation generally resulted in significantly
higher enzyme activities than the liquid cultures. Therefore, two solid state fermentation
systems were used in the present assays: mixture of chopped corn stalks and corn leaves, and
their mixtures with wheat bran at a ratio of 1:1. Fermentations were carried out for 12 days at
25 °C or 37 °C, and enzyme activities were determined from the crude water extracts obtained
every second day of the cultivation. Isolates grew intensively on these substrates, and high
activities of the cellulolytic enzymes and lipase have been observed during the fermentation
period. The total cellulolytic activity of the crude extracts was determined by using Whatman
No. 1 filter paper as substrate; in case of each isolate, the highest amount of reducing sugar
was measured at the second day of the fermentation. It is worth to point out that significantly
higher total cellulolytic activity was usually detected in the crude extracts derived from
fermentation medium containing wheat bran. It might be due to the fact that wheat bran
supplied convenient amount of nutrients and porosity for oxygen supply. In both fermentation
systems, cellobiohydrolase activity of the tested fungi was found to be significantly lower than
their endoglucanase and β-glucosidase activities; additionally, it was detected that the activity
of endoglucanase generally reaches its maximum during the first half of the incubation, while
β-glucosidase on the sixth day or later. Among the tested representatives of the
abovementioned genera, a Rhizomucor sp. and a Rhizopus sp. strains proved to be outstanding
in its lipase, and protease producing ability, respectively. The investigated fungi could
potentially be applicable for biodegradation of agro-industrial wastes and efficient production
of industrial enzymes on these cheap, easily available substrates.