Phosphoric Acid-Assisted Enzymatic Production of Water-Soluble Cellulosic Oligomers

Water-soluble cellulosic oligomers (WCOs) are increasingly recognized for their prebiotic benefits, but their efficient enzymatic production is hindered by the high crystallinity of cellulose, which limits enzyme accessibility. This study introduced an efficient and scalable strategy combining phosphoric acid pretreatment with enzymatic hydrolysis to produce high-purity WCOs. Microcrystalline cellulose treated with 85 wt% phosphoric acid at 10 °C exhibited significantly reduced crystallinity and crystallite size, improving its susceptibility to enzymatic degradation. Subsequent hydrolysis of the hydrated regenerated cellulose (HRC85-10) using Celluclast® at pH 7.0 for 1 h resulted in a WCO selectivity of 93.5%, with cellobiose and cellotriose identified as major oligomeric products via electrospray ionization mass spectrometry. Maintaining cellulose in a hydrated form significantly improved both the yield and selectivity of WCOs. In vivo studies further confirmed the prebiotic potential, with a significant increase in fecal Lactobacillus spp. and Bifidobacterium spp. (p < 0.05) following WCO supplementation. These findings demonstrated a practical and effective approach for producing functional WCOs for use in dietary and gut health applications.