# Sustainable Endoglucanase Production from Lignocellulosic Waste Through Fungal Co-Culture Technology: A Step Towards Circular Economy

## TL;DR

This paper presents a sustainable method to produce endoglucanase from agricultural waste using a fungal co-culture, promoting circular economy and bioeconomy concepts.

## Contribution

The novel use of a fungal co-culture for enzyme production from pea hulls supports waste valorization and circular bioeconomy.

## Key findings

- Alkaline-pretreated pea hulls were the most effective substrate for endoglucanase production.
- Optimized conditions achieved a maximum endoglucanase activity of 119.58 U/mL/min.
- SEM analysis showed structural disruption in biomass, enhancing enzyme accessibility.

## Abstract

Large amounts of agricultural waste rich in lignocellulose are produced every year and often remain underutilized. This study demonstrates a sustainable way to convert such waste into a valuable industrial enzyme, endoglucanase, using a fungal co-culture of Aspergillus fumigatus and Rhizopus arrhizus. Among several tested residues, alkaline-pretreated pea hulls proved to be the most effective substrate due to their high cellulose and low lignin content after treatment. Process conditions were statistically optimized using Design of Experiments approaches, which significantly enhanced enzyme production. The optimized enzyme system was further applied to hydrolyze pretreated biomass, releasing a high amount of fermentable sugars. These findings highlight the dual role of pea hulls as both a substrate for enzyme production and a feedstock for biomass hydrolysis, supporting waste valorization and circular bioeconomy concepts.

This study focused on optimizing endoglucanase production using a peculiar fungal co-culture comprising Rhizopus arrhizus and Aspergillus fumigatus, identified through morphological and 18S rDNA analyses. The co-culture achieved the highest enzyme production after 72 h of fermentation with alkaline-treated substrates. Scanning Electron Microscopy (SEM) revealed substantial structural disruption in pretreated biomass, enhancing enzyme accessibility. Among the tested substrates, pea hulls proved to be the most effective for enzyme production. Optimization of physical and nutritional parameters was performed using Design of Experiments (DOE) approaches, specifically Plackett–Burman Design (PBD) for screening and Central Composite Design (CCD) for fine optimization. The maximum endoglucanase activity of 119.58 U/mL/min was obtained under the optimized conditions of 27.5 °C, pH 5.5, inoculum age 3.5 days, and supplementation with 1.5% fructose, 1.25% yeast extract, 1.25% sodium nitrate, and 1.25% Tween 80. Analysis of Variance (ANOVA) confirmed the significance of these parameters and their interactions at a 95% confidence level, with a strong model fit (R2 = 0.9052). This study demonstrates the potential of waste pea hulls as a cost-effective substrate for enzyme production, supporting waste valorization and contributing to a circular bioeconomy through sustainable biomass utilization.