Clavulanic acid (CA), a potent beta-lactamase inhibitor, is produced by a filamentous bacterium. Here, the effect of DO and shear, expressed as impeller tip velocity, on CA production was examined. Cultivations were performed in a 4 L fermentor with speeds of 600, 800 and 1,000 rpm and a fixed air flow rate (0.5 vvm). Also, cultivation with automatic control of dissolved oxygen, at 50% air saturation, by varying stirrer speed and using a mixture of air and O(2) (10% v/v) in the inlet gas, and a cultivation with fixed stirrer speed of 800 rpm and air flow rate of 0.5 vvm, enriched with 10% v/v O(2), were performed. Significant variations in CA titer, CA production rate and O(2) uptake-rate were observed. It was also found that the DO level has no remarkable effect on CA production once a critical level is surpassed. The most significant improvement in CA production was related to high stirrer speeds.

The development of process engineering approaches to integrate the production of biofuels and high value-added biobased products, such as enzymes and nanocellulose, is crucial to improve the financial performance and sustainability of lignocellulosic biomass biorefineries. Here, the feasibility of applying enzymes produced on-site to obtain nanocellulose was evaluated using eucalyptus cellulose pulp as a model feedstock. A systematic analysis of the structural properties of the nanomaterials obtained after hydrolysis using a cellulolytic enzymatic complex with high endoglucanase specific activity (17.09 IU/mg protein ), produced by Aspergillus niger, followed by sonication, revealed that longer ball milling pretreatment and reaction times favored extraction of the cellulose nanocrystals (CNCs). The highest yield (24.6%) of CNCs was achieved using 96 h of enzymatic hydrolysis of the ball-milled cellulose pulp, followed by sonication for 5 min. The CNCs presented approximate lengths of 294.0 nm and diameters of 24.0 nm, and the crystallinity index increased from 57.5% to 78.3%, compared to the cellulose pulp that was only ball milled. These findings demonstrated that nanocelluloses could be successfully extracted using on-site produced enzymes and that the sustainable integrated process reported here could contribute to the development of the nascent biobased economy.

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Sequential solid-state and submerged cultivation with sugarcane bagasse as substrate for cellulase production by Aspergillus niger A12 was assessed by measuring endoglucanase activity. An unconventional pre-culture with an initial fungal growth phase under solid-state cultivation was followed by a transition to submerged fermentation by adding the liquid culture medium to the mycelium grown on solid substrate. For comparison, control experiments were conducted using conventional submerged cultivation. The cultures were carried out in shake flasks and in a 5-L bubble column bioreactor. An endoglucanase productivity of 57 ± 13 IU/L/h was achieved in bubble column cultivations prepared using the new method, representing an approximately 3-fold improvement compared to conventional submerged fermentation. Therefore, the methodology proposed here of a sequential fermentation process offers a promising alternative for cellulase production.

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