Mixed Cultures of Aspergillus niger and Rhizopus oryzae Using Lignocellulosic Substrates to Improve Hydrolytic Enzyme Production

This study aims to evaluate the potential of filamentous fungi consortiums to design a bioprocess for producing hydrolytic enzymatic cocktails on lignocellulosic biomass. Four different microorganisms were analyzed in solid and submerged monoculture fermentation systems: Aspergillus niger, Trichoderma harzianum, Rhizopus oryzae, and Thermomyces lanuginosus. A. niger and R. oryzae were selected as the most suitable to form a consortium. Two different inoculation strategies were assessed, one was simultaneous and the other was a 12-h post-phase inoculation. Xylanase, cellulase, amylase, pectinase, and protease activities were evaluated in mono and mixed cultures. Furthermore, the secretomes of the fermentation systems were analyzed by mass spectrometry to compare and determine the difference in the protein profiles excreted. The mixed culture of A. niger and R. oryzae, which were inoculated simultaneously in solid-state fermentation, showed the highest levels of enzyme at 96 h, 30°C, 1 × 10⁷ conidia, and 65% relative humidity. The proportion of xylanase (98 IU/gds), cellulase (27 IU/gds), amylase (30 IU/gds), pectinase (21 IU/gds) and protease (108,000 IU/gds) was enhanced by 25% regarding the monoculture, demonstrating that the use of microbial consortiums can be a promising alternative to obtain enzymatic cocktails with high synergism.