Multi-Step Pretreatment Strategies for Enhancing Cellulase Accessibility in Agro-Residues

Abstract

Lignocellulosic biomass, especially agro-industrial residues such as sugarcane bagasse, represents a plentiful and sustainable resource for bioconversion-based value generation. However, the inherent structural complexity and recalcitrance of lignocellulose significantly restrict enzymatic hydrolysis and limit overall bioprocess efficiency. In recent years, multi-step pretreatment strategies have gained considerable attention as effective approaches to enhance biomass digestibility through sequential and targeted modification of its physicochemical characteristics. This review provides a comprehensive evaluation of the underlying principles, mechanisms, and performance of multi-step pretreatment methods, with particular emphasis on improving cellulase accessibility. Combinations of physical, chemical, physicochemical, and biological pretreatments are critically analyzed in terms of their influence on lignin disruption, hemicellulose removal, reduction in cellulose crystallinity, enhancement of porosity, and improvement in hydrolytic efficiency. In addition, key challenges associated with process integration, techno-economic feasibility, and environmental sustainability are discussed, along with future perspectives for the development of efficient and scalable lignocellulosic biorefineries.