Abstract
INTRODUCTION: Laccase exhibits significant applications in food additives, wastewater treatment, and biomass processing. Fungal laccase exhibits high activity, good stability, and excellent performance. However, scale-up production of high-yield laccase from fungi is challenging. This study aimed to identify crucial factors affecting enzyme production and analyze the enzymatic properties of laccase during fermentation. METHODS: A laccase-producing white-rot fungus was used for fermentation process optimization in 200 L and 1200 L fermenters. The Plackett-Burman design revealed three significant influencing factors: temperature, aeration ratio, and agitation speed. The steepest ascent experiment was used to approximate the maximum response region, followed by the establishment of a regression model between experimental factors and laccase activity using the Box-Behnken response surface methodology and optimal fermentation condition selection. RESULTS: The optimal conditions for laccase production by Ganoderma lucidum fermentation were 30°C temperature, 0.66 aeration ratio, and 100 rpm agitation speed, achieving a maximum laccase activity of 214,185.2 U/L. Dissolved oxygen (DO) was a crucial factor for high laccase yield, and its maintenance at a high level in the fermentation system significantly enhanced the enzyme activity. Fermentation batches with varying laccase production levels exhibited a trend of decreasing pH and a subsequent increase in the mid-to-late fermentation stages. With decreased pH, the DO level started declining; when DO stabilized, the pH started to rebound, coinciding with the peak laccase activity, indicating a signal of fermentation endpoint in industrial production. DISCUSSION: This study provides valuable theoretical and data support for the industrial production of laccase by fungi through optimized fermentation processes.