# Eucalyptus lignin production coupled with Cellic ® CTec3 HS enzymatic hydrolysis to monosaccharides

## TL;DR

This paper presents a two-step process to extract lignin from eucalyptus and convert the remaining material into high concentrations of glucose and xylose.

## Contribution

A new hydro-organo-thermal method using diluted imidazole and optimized enzymatic hydrolysis for high sugar yields.

## Key findings

- 70 wt% lignin removal was achieved under optimized conditions.
- Glucose and xylose yields reached 99 and 92 mol%, respectively.
- A total reducing sugar concentration of 91 g/L was obtained in the hydrolysate.

## Abstract

The aim of this work was to design, develop, and optimise a two-step process for extracting lignin from Eucalyptus globulus residues and subsequently converting the remaining holocellulose into monosaccharides, primarily glucose and xylose. Firstly, the solubility of high-quality lignin was optimised using a new hydro-organo-thermal method involving a diluted imidazole solution in water instead of pure imidazole. Subsequently, enzymatic hydrolysis of the polysaccharides-rich pretreated solid was optimised to give glucose and xylose by using the commercial enzymatic mixture Cellic® CTec3 HS (Novozymes, Bagsværd, Denmark) supplemented with endo-1,4-β-xylanase from Trichoderma viride (Merck, Darmstadt, Germany). The extraction process was thoroughly optimised by acting on several parameters, such as the imidazole concentration, temperature, reaction time, and solid-to-liquid ratio. Under the optimised reaction conditions (imidazole 35 wt%, biomass loading 10 wt%, 120 °C, 2 h, 400 rpm) over the entire process, about 70 wt% of lignin removal was achieved, together with glucose and xylose yields of 99 and 92 mol%, respectively, reaching a high total sugar concentration of 91 g/L in the hydrolysate.

• A new hydro-organo-thermal treatment of lignocellulosic biomass has been proposed.

• Glucose and xylose yields were 99 and 92 mol% in 48 h at low enzyme dosage.

• A total reducing sugar concentration of 91 g/L was achieved in the hydrolysate.

The online version contains supplementary material available at 10.1007/s00253-026-13793-2.