Adsorption and desorption of cellulase on/from lignin pretreated by dilute acid with different severities

https://doi.org/10.1016/j.indcrop.2020.112309Get rights and content

Highlights

  • Adsorption/desorption of cellulase on enzymatic residual lignin (ERL) was studied.

  • Dilute acid (DA) pretreatment of ERL at higher severity intensified the adsorption.

  • Desorption capacity of cellulase from ERL was decreased by DA pretreatment.

  • Recoveries of cellulase from ERLs decreased with the increase of severity factors.

  • Bound-cellulase on DA pretreated ERLs exhibited activity, but was hard to be reused.

Abstract

Desorption of cellulase from lignin is necessary to recycle cellulase in enzymatic hydrolysis. Currently, the knowledge about cellulase desorption from lignin is still limited. In present work, the adsorption and desorption of cellulase on/from enzymatic residual lignins (ERLs) pretreated with dilute acid at different severities were investigated. The results showed that the molecular weights of dilute acid-pretreated ERLs (DA-ERLs) decreased with the increase of severity. The hydrophobicity and contents of condensed phenolic structures in ERLs increased after DA pretreatment with the increase of severity. Compared with un-pretreated ERL, DA-ERLs pretreated with severity factors of 1.17, 1.51, and 1.69 increased the inhibition degree of cellulose hydrolysis from 21.5 % (ERL) to 24.6 %, 28.5 % and 31.9 %, respectively. DA-ERLs pretreated at higher severities exhibited larger adsorption capacities with individual cellulase components. DA pretreatment of ERL at higher severities intensified the binding with cellulase. The binding strength between lignin and cellulase increased from 236.0 mL/g to 261.5–410.9 mL/g with the increase of severity. DA pretreatment decreased the desorption capacity of cellulase from ERLs. The recovery of cellulase from lignin decreased from 78.6% to 36.9%–14.7 % with the increase of severity. The cellulase desorbed from ERLs remained portion of activities, but the cellulase bound on DA-ERLs was hard to be released and provided a lower glucose yield (4.8 %–5.9 %) than that on un-pretreated ERL (16.2 %). The results reported here will aid cellulase recycling and efficient hydrolysis of lignocellulose after dilute acid pretreatment.

Introduction

Lignocellulosic biomass is viewed as a potential alternative to fossil fuels to reduce environment pollution. The development of biorefinery is a revolutionary concept to convert waste to resources such as the production of clean energy, paper, and chemicals (Himmel et al., 2007; Kamali et al., 2019; Mandeep et al., 2019, 2020; Yiin et al., 2019). Enzyme, as a biocatalyst with high catalytic activity and substrate-specificity, has excellent performance in industrial processes for high product purity, fewer side reactions, and environmental friendliness (Kumar et al., 2018, 2019). The recalcitrance of lignocellulosics is considered as a barrier in economically achieving release of sugars for producing biofuels and bio-based chemical production (Himmel et al., 2007). Pretreatment is crucial to break the recalcitrance of feedstock for its efficient enzymatic hydrolysis (Galbe and Zacchi, 2012). Among many pretreatments, dilute acid (DA) pretreatment was one of the efficient techniques for overcoming biomass recalcitrance (Huang et al., 2019; Loow et al., 2016; Qin et al., 2019; Sipponen et al., 2017). DA pretreatment increased the cellulose accessibility by hemicellulose removal and lignin redistribution, whereas the lignin content increased after DA pretreatment (Jensen et al., 2017; Meng et al., 2015). After DA pretreatment, the morphology and structure of lignin changed significantly (Sipponen et al., 2017), and lignin was degraded more severely with increasing acid concentration (Jensen et al., 2017). Furthermore, the lignin remained in the pretreated lignocellulose exhibited stronger inhibitory effect after severe pretreatment with acid as catalyst (Kellock et al., 2019).

It was extensively reported that residual lignin in lignocellulose after pretreatment inhibited enzymatic hydrolysis by physical hindrance and non-productive adsorption with cellulase (Hao et al., 2019; Rahikainen et al., 2011; Selig et al., 2007). Although Djajadi et al. (2018) reported that the retardation of cellulose degradation was due to the physical hindrance by lignin rather than non-productive adsorption of cellulase by lignin, considerable evidences indicated that binding of cellulase non-productively onto lignin was considered as the main reason for the inhibition and residual lignins from different pretreatments exhibited different lignin-cellulase binding properties (Huang et al., 2017; Kellock et al., 2019; Nakagame et al., 2011; Yang and Pan, 2015). Currently, the knowledge about the effect of DA pretreatment severity on enzyme adsorption onto lignin is quite limited.

Recycling enzymes from solid residues after hydrolysis is a potential strategy to decrease enzyme dosage in bioconversion of biomass (Weiss et al., 2013; Xin et al., 2020). As reported, a large fraction of cellulase can bind onto the gradually accumulated lignin in enzymatic hydrolysis and thus it is necessary to maximize enzyme amount desorbed from lignin (Hao et al., 2019; Rahikainen et al., 2011; Tu et al., 2007). The cellulase adsorbed on lignin can be recovered by washing with fresh buffer or sometimes with the addition of surfactants (Li et al., 2016). Kumar and Wyman (2009) reported that the cellulase adsorbed on poplar enzymatic residual lignin after DA pretreatment was completely removed by fresh buffer replacement. Aside from the utilization of cellulase by desorption, the residual lignin could also be collected and added into subsequent hydrolysis to recycle enzyme directly (Tu et al., 2007). However, various studies have shown that the desorption of enzyme from lignin is not easy and partially irreversible (Kellock et al., 2019; Qi et al., 2011; Rahikainen et al., 2011). At present, the effect of DA pretreatment severity on enzyme desorption from residual lignin was still not clarified.

In this work, enzymatic residual lignin (ERL) was treated with dilute sulfuric acid at different severities to investigate the inhibitory effect on cellulose digestion. Typically, DA pretreatment was performed at temperatures from 120 to 210 °C, acid concentration of less than 4 wt%, and with residence time of several minutes to an hour (Xu and Huang, 2014). Baadhe et al. (2014) reported a higher concentration of fermentable sugars from DA pretreated corncob after pretreatment with 0.25 M (2.4 wt%) H2SO4 at 121 °C for 20 min. Therefore, the ERL isolated from corncob was pretreated with 0.5 %, 1.0 %, and 1.5 % (w/w) sulfuric acid at 120 °C for 1 h, respectively. The adsorption and desorption properties of cellulase on the ERLs and the hydrolytic capacity of cellulase desorbed from ERLs were investigated in detail. The results would provide guidance to recycling cellulase from residual lignin and development of DA pretreatment for efficient hydrolysis of lignocelluloses.

Section snippets

Materials

ERL can represent the original lignin in plant cell wall due to its minimal structural changes (Hao et al., 2019; Rahikainen et al., 2011). The ERL from corncob used in this work was purchased from Longlive Bio-Technology Co., Ltd. (Shandong, China). Chemical composition of ERL was determined as the method in the NREL protocol (Sluiter et al., 2012). The ERL contained 93.5 wt% Klason lignin and 2.3 wt% acid-soluble lignin. Microcrystalline cellulose (Avicel PH 101), bovine serum albumin (BSA), p

Characterization of ERLs

It was reported that the severity factor combined with pH value was better suited to estimate the intensity of DA pretreatment (Chum et al., 1990). In this work, the severity factors of DA pretreatment increased from 1.17 to 1.69 when the acid concentration increased from 0.5 % (w/w) to 1.5 % (w/w). Lignin characteristics were considered to be main factors for the non-productive binding of cellulase and negative effect on cellulose digestibility (Cheng et al., 2015; Sipponen et al., 2017). To

Conclusions

ERL was deconstructed more severely and generated greater amounts of condensed structures after DA pretreatment with the increase of severity. ERLs became more porous and hydrophobic after DA pretreatment with higher severities. DA pretreatment of ERL at higher severities intensified the binding between ERLs and cellulase and caused stronger inhibitory effects. More individual cellulase components were bound on ERLs with the increase of severity, and it would be of interest to investigate the

CRediT authorship contribution statement

Jinye Wang: Investigation, Methodology, Writing - original draft. Xixun Hao: Methodology, Formal analysis. Peiyao Wen: Methodology, Formal analysis. Tian Zhang: Resources, Validation. Junhua Zhang: Conceptualization, Writing - review & editing, Project administration.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (Nos. 31670598, 31270622).

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