Subsequent Enzymatic Saccharification Research Articles

Upgrading lignocellulosic biomass to high-value products through the pretreatment driven by bio-based ethylene glycol solvent

Sustainable processing of lignocellulosic biomass is meaningful for comprehensive biomass conversion. Here, a ball milling (BM) assisted biomass-derived ethylene glycol (BEG) pretreatment is proposed to removes nearly 90 % of the lignin and increases cellulose accessibility by 3 times than raw biomass, facilitating subsequent enzymatic saccharification or catalytic conversion for 5-hydroxymethylfurfural (HMF), achieving yields of 92.9 % and 32.6 %. The synergistic effect of BM/BEG positively impacted delignification, followed by saccharification and HMF conversion. Introduction of biomass-derived solvent recovers the lignin with a well-preserved chemical structure, uniform molecular weight, short side chains, and high purity, making it highly suitable for downstream utilization. Density functional theory revealed hydrogen bonding and good miscibility between BEG and lignin units, suggesting reaction mechanisms and pathways. Molecular dynamics simulations showed that the solubilizing effect of BEG on lignin promoted cellulase-cellulose binding, thereby enhancing saccharification efficiency. The successful use of BEG solvents to maximize the biomass value is conducive to achieve closed-loop biorefinery approaches.

In situ pretreatment of wood samples with deep eutectic solvents for enhanced lignin removal and enzymatic Saccharification efficiency optimization

Deep eutectic solvents (DESs) are commonly utilized as an effective pretreatment method for biomass materials, particularly for enhancing the value of lignocellulosic biomass. However, to the best of our knowledge, the utilization of DESs for in situ pretreatment of wood samples has been seldom reported. In this study, the ability of five DESs systems to in-situ pretreatment of balsa wood samples was investigated. Among of them, the Choline Chloride-Ethylene Glycol-P-Toluenesulfonic Acid (ChCl-EG-PTSA) system showing an excellent lignin removal rate (the lignin removal of 87.34 % and cellulose retention of 86.32 %), which is significantly better than the delignification effect of the traditional acid chlorite method (lignin removal of 72,13 % and cellulose retention of 64.57 %). In addition, in the subsequent enzymatic saccharification experiments, after the wood samples were pretreated in situ by the ChCl-EG-PTSA system, the enzymatic saccharification yield of cellulose reached 75.46 %, and the enzymatic saccharification yield of xylose reached 92 %. At the same time, this study also used SEM, FTIR, XPS, TGA and other testing technologies to conduct detailed analysis and characterization of the pretreated samples. This comprehensive analysis unequivocally demonstrated the viability of the ChCl-EG-PTSA system as an environmentally sustainable and efficient approach for pretreating forest biomass. Significantly, this in-situ wood pretreatment strategy significantly lays a robust research groundwork for the development of multifunctional wood fiber composites in subsequent stages.

Unlocking the potential of spent coffee grounds via a comprehensive biorefinery approach: production of microbial oil and carotenoids under fed-batch fermentation.

The valorization of renewable feedstock to produce a plethora of value-added products could promote the transition towards a circular bioeconomy. This study presents the development of cascade processes to bioconvert spent coffee grounds (SCGs) into microbial oil and carotenoids employing sustainable practices. The stepwise recovery of crude phenolic extract and coffee oil was carried out using green or recyclable solvents, i.e., aqueous ethanol and hexane. Palmitic acid (43.3%) and linoleic acid (38.9%) were the major fatty acids in the oil fraction of SCGs. The LC-MS analysis of crude phenolic extracts revealed that chlorogenic acid dominated (45.7%), while neochlorogenic acid was also detected in substantial amounts (24.0%). SCGs free of coffee oil and phenolic compounds were subjected to microwave-assisted pretreatment under different irradiations and solvents to enhance subsequent enzymatic saccharification. Microwave/water pretreatment at 400 W, followed by enzymatic hydrolysis with proteases, hemicellulases, and cellulases, at 50g/L initial SCGs, led to satisfying overall yields of cellulose (75.4%), hemicellulose (50.3%), and holocellulose (55.3%). Mannan was the most extractable polysaccharide followed by galactan and arabinan. SCGs hydrolysate was used in fed-batch bioreactor fermentations with Rhodosporidium toruloides to produce 24.0g/L microbial oil and carotenoids of 432.9μg/g biomass.

Revealing the mechanisms of lignin modification by phenolic additives during pre-hydrolysis treatment and its effects on enzymatic hydrolysis

Condensed lignin formed during eucalyptus woodchip pre-hydrolysis adversely affects subsequent enzymatic saccharification, and phenolic additives are popular lignin condensation inhibitors. The goal of this project was to reveal the mechanisms of lignin modification by phenolic additives during woodchip pre-hydrolysis and its effects on subsequent enzymatic saccharification. The HSQC and 31P NMR analyses confirmed that resorcinol with three electrophilic substitution of active sites could act as a cross-linking agent to promote the formation of condensed lignin, whereas 2-naphthol and sodium 2-naphthol-7-sulfonate could block benzyl carbocations and reduce the formation of condensed lignin. Interestingly, the introduction of sulfonic acid groups increased the electrostatic repulsion between enzyme and lignin as well as increased the hydrophilicity of lignin, thereby greatly reducing non-productive adsorption and physical obstacle effects. Sodium 2-naphthol-7-sulfonate as an efficient benzyl carbocations scavenger reduced the non-productive adsorption and physical obstacle effects of lignin by 82.4% and 13.3%, respectively.

Ultrasound treatment of herbal extraction residue to enhance enzymatic saccharification

The utilization of herbal extraction residues (HERs) for production of valuable products has gained significant momentum in recent years. In this study, Lianhua Qingwen residue (LQR) was selected as a representative HER for enzymatic saccharification. By subjecting starch-containing raw unpretreated LQR to direct hydrolysis using a combination of cellulolytic and amylolytic enzyme, a glucan conversion of around 60 % was achieved within 48 h. To further enhance the hydrolysis rate and yield, ultrasound treatment of LQR samples was conducted. Investigation into the effect of ultrasound treatment on subsequent enzymatic saccharification revealed a noteworthy acceleration in the saccharification rate of LQR, exceeding 20 % within 10 h of hydrolysis time. Notably, this enhancement was achieved even when the substrate concentration was high at 10 % under optimized conditions of 5 min of sonication at a power intensity of 1.20 kW/cm2 and a temperature of 50 °C. The improvement in efficiency can be attributed to the disruptive effect of ultrasound treatment on the structure of LQR, leading to a reduction in particle size. Consequently, the LQR particles exhibited better contact with the enzyme, promoting a more efficient saccharification process. These findings suggest that ultrasound technology holds considerable potential for enhancing the enzymatic saccharification of HERs.

Long term study of cell wall saccharification and high added value phenolic acids production from Phalaris aquatica L. energy crop

Phalaris aquatica L. is a cool-season perennial grass of the Mediterranean that has been used for forage production, while recently was identified as a novel candidate for sustainable second-generation bioethanol production. This study provides for the first time evidence about this energy crop long-term performance (five years of growth), the cell wall saccharification efficiency and the production of high added value compounds during this period. High biomass production was recorded constantly after the first year of plant growth, with a maximum of 22.4 ± 0.5 t of dry mass (DM) per ha and stable structural polysaccharides concentration 68.8% ± 0.8 w/w DM from the third year on. Independently from crop age, dilute acid pretreatment (120 °C, 1.5% H2SO4, 45 min) promoted the subsequent enzymatic hydrolysis of P. aquatica L. biomass. The dilute acid pre-treatment positive effect is attributed both to the hemicellulose solubilization and the breakage of intermolecular bonds between polysaccharides and phenolic acids, as supported by the present study results These combined acid/enzymatic hydrolysis processes were able to convert more than 80% of cell wall polysaccharides into fermentable sugars, releasing simultaneously a substantial amount of phenolic acids of 2.16% ± 0.48 w/w DM. The ratio of ferulic to p-coumaric acid in the acid hydrolysate can be used for the prediction of the subsequent cellulose enzymatic saccharification efficiency, by the model described here. The protocatechuic acid, which can find many technological applications, was the main bioactive polyphenol in the final hydrolysate with concentration ∼1% w/w DM. These results are supportive of Phalaris aquatica L. suitability as a sustainable biorefinery crop, under the semi-arid Mediterranean climatic conditions contributing to this region biodiversity and prosperity.

Acid/alkali-catalyzed deep eutectic solvent pretreatment of corn straw for enhanced biohydrogen production

Lignocellulose is considered as a potential source for biohydrogen production, however, the tough and rigid structure endow the lignin with high resistance to biodegradation. In this work, choline chloride/1,4-butanediol deep eutectic solvent (DES) pretreatment aiding with AlCl3/NaOH was developed to dissociate the lignin macromolecular structure of corn straw for promoting biohydrogen production. The results showed that alkali-catalyzed DES pretreatment reached a yield of biohydrogen production of 117.4 mL/g total solids (TS) in comparison with that of acid-catalyzed DES pretreatment (100.5 mL/g TS) at 100 °C for 3 h with 4.0 wt% alkali/acid dosage. The enhanced biohydrogen production was mainly ascribed to the higher lignin removal rate (81.3%) of alkali-catalyzed DES pretreatment, which facilitated subsequent enzymatic saccharification and dark fermentation of carbohydrate. Moreover, the recovered lignin obtained from this pretreatment displayed well-preserved β-O-4′ linkages (44.4/100Ar) with an appropriate molecular weight (1540 g/mol) and regular spherical particles. The new insights of this work may provide guidance for designing novel DES pretreatment toward bio-energy production from lignocellulose fractionation and component valorization.

Biotechnological enhancement of lactic acid conversion from pretreated palm kernel cake hydrolysate by Actinobacillus succinogenes 130Z

The aim of this study was to establish an improved pretreatment and fermentation method i.e. immobilized cells for high recovery of fermentable sugars from palm kernel cake (PKC) and its effects on fermentability performance by Actinobacillus succinogenes 130Z in the conversion of the fermentable sugar to lactic acid. The effects of oxalic acid concentrations (1–6% w/v) and residence times (1–5 h) on the sugar recovery were initially investigated and it was found that the highest mannose concentration was 25.1 g/L at the optimum hydrolysis conditions of 4 h and 3% (w/v) oxalic acid. The subsequent enzymatic saccharification of the pretreated PKC afforded the highest enzymatic digestibility with the recovered sugars amounting to 25.18 g/L and 9.14 g/L of mannose and glucose, respectively. Subsequently, the fermentability performance of PKC hydrolysate was evaluated and compared in terms of cultivation phases (i.e. mono and dual-phases), carbonate loadings (i.e. magnesium and sodium carbonates), and types of sugars (i.e. glucose and mannose). The highest titer of 19.4 g/L lactic acid was obtained from the fermentation involving A. succinogenes 130Z in dual-phase cultivation supplemented with 30 g/L of magnesium carbonate. Lactic acid production was further enhanced by using immobilized cells with coconut shell-activated carbon (CSAC) of different sizes (A, B, C, and D) in the repeated batch cultivation of dual-phase fermentation producing 31.64 g/L of lactic acid. This work sheds light on the possibilities to enhance the utilization of PKC for lactic acid production via immobilized A. succinogenes 130Z.

Acid and alkali pretreatment of corn stalk for fractal‐like kinetic analysis of enzymatic saccharification

AbstractBACKGROUNDThe conversion of lignocellulose to sugar is part of the process of lignocellulose biorefining and, therefore, is crucial for effective lignocellulose utilization.RESULTSIn this study, alkali and acidic pretreatments were carried out to deconstruct corn stalk (CS) material for efficient subsequent enzymatic saccharification. Most xylan and lignin can be removed from CS material by acid and alkali pretreatments, respectively, as verified by XRD (X‐ray diffractometer) and FT‐IR (Fourier Transform Infrared Spectroscopy) analyses. Fractal‐like kinetics were assessed to analyze the effect of enzyme loading on the glucose and xylose sugar yields from the two pretreated CS materials, with fractal dimension considered as a dominant factor in rate constant decrease. Fractal dimension analysis demonstrated that the alkali pretreated CS was more easily hydrolyzed that the acid pretreated CS. For glucan hydrolysis, the effect of enzyme loading on the rate constant of the alkali pretreated CS was marginally higher than that of the acid pretreated CS. For xylan hydrolysis, enzyme loading was shown to significantly improve the rate constant of the alkali pretreated CS compared to the acid pretreated CS. Furthermore, the developed fractal‐like model exhibits high prediction accuracy for sugar yield at high enzyme loadings.CONCLUSIONThe application of the fractal‐like theory was quite successful for studying enzymatic kinetics. © 2022 Society of Chemical Industry (SCI).

Alkali-facilitated deep eutectic solvent for effective bamboo saccharification

Herein, a Na2S promoted deep eutectic solvent (DES) was established to reduce the natural recalcitrance of moso bamboo (MB) and improve the subsequent enzymatic saccharification. It was found that the addition of Na2S (Choline chloride/Ethylene glycol/Na2S) dramatically promoted the deconstructions of lignin with highest removal of 74.67 %, but at the same time preserved glucan and hemicellulose to the maximum extent. With the fractionation, the enzymatic saccharification yield of pretreated MB can reach 100 % under the pretreatment condition of 140 °C, and lignin could be readily recovered with a high yield of 81.47 %. The proposed DES is superior to normal alkaline DES in terms of the higher lignin removal and recovery yield, carbohydrate preservation and enzymatic digestibility, which indicated Na2S as a novel and powerful reinforcer enhancing the DES fractionation efficiency.

The effect of soluble phenolic compounds from hydrothermally pretreated wheat straw on Trichoderma reesei cellulases and commercial enzyme cocktails

Hydrothermal methods are commonly applied in pretreatment of lignocellulose for conversion to biofuels and chemicals. This pretreatment partially disassembles and solubilizes hemicelluloses and lignin, releasing sugars and phenolic compounds, which are potential inhibitors for the subsequent enzymatic saccharification step. In this work, the inhibitory effect of soluble phenolics in hydrothermal pretreatment liquor on canonical Trichoderma reesei enzymes was investigated, using purified cellobiohydrolases, endoglucanases, a xylanase, and a lytic polysaccharide monooxygenase, as well as commercial enzyme cocktails Cellic Ctec2 and Celluclast. The pretreatment liquor was fractionated to enrich the phenolics, and thoroughly analyzed. The most sensitive to phenolics inhibition were cellobiohydrolases, the major enzymes in crystalline cellulose degradation and the primary component in cellulolytic mixtures. These were inhibited by the oligophenolics and phenol-carbohydrate complexes (lignin-carbohydrate complex type of compounds), the latter of which could be mitigated by other enzymes, i.e., xylanases and endoglucanases. Addition of lytic polysaccharide monooxygenase to the hydrolysis reactions was found to relieve the negative effect of phenolics. Similarly, hydrolytic activity of a commercial enzyme cocktail Cellic Ctec2 was enhanced in the presence of low concentrations of phenolics.

Whole slurry saccharification of mild oxalic acid-pretreated oil palm trunk biomass improves succinic acid production

The present work compared two different routes in preparing oil palm trunk biomass (OPTB) hydrolysates for producing industrially-valuable succinic acid (SA). OPTB was initially pretreated with mild oxalic acid (1% w/v) under different reaction time to investigate its effect on the total sugar and inhibitor yields during pretreatment and subsequent enzymatic saccharification. Substantial solubilisation of hemicellulose (56–59 %) was achieved after pretreatment for 3−4 h. After enzymatic saccharification, high cellulose digestibility of 86–87 %, with 22.1–22.2 g sugar per 100 g of raw OPTB from washed pretreated solids (Route 1), and 71–74 %, with 43.2–44.9 g sugar per 100 g of raw OPTB from whole slurry (Route 2) saccharification, respectively was achieved. Fermentation of the hydrolysates derived from Route 2 by Actinobacillus succinogenes 130Z afforded a twofold increase in the SA production compared to that from Route 1. Mass balance calculations showed that 175 g of SA could be produced per kg of OPTB via Route 2 vs. 153 g via Route 1. The proposed approach in this study offers pioneering insights into the greatest improvement in SA yield achievable via efficient biomass utilisation.

Improved sugar recovery from enzymatic hydrolysis of Miscanthus sinensis by surfactant-mediated alkaline pretreatment

Surfactants play a vital role in the removal of lignin and subsequent enzymatic saccharification of lignocellulosic biomass. In this work, a surfactant-mediated alkaline pretreatment (SMAP) with short residence time and low NaOH loading has been successfully developed for enhancing enzymatic hydrolysis of a potential energy plant, Miscanthus sinensis, and evaluating its potential as a feedstock for biofuel production. As compared to individual alkaline pretreatment (AP), the SMAP process resulted in significantly more hemicellulose removal, more efficient delignification, higher cellulose content (55% vs. 51%), and stronger morphological disruption of the pretreated stalks, making them more accessible to cellulases and/or microorganisms. As a result, SMAP produced a significantly higher sugar yield of 402.5 mg/g than the control and AP group (287.8 mg/g) (p < 0.05). Among the tested surfactants, Tween 40 was identified as a potential candidate for SMAP of biomass wastes. SMAP with mild conditions (i.e., 121 °C, 10 min, 1.0% NaOH, 1.0% Tween 40) gave the highest sugar yield of 566.0 mg/g, which were 6.5-folds greater than the raw stalk. These demonstrations for using SMAP to achieve enhanced lignin removal and sugar recovery from biomass highlight its strong application potentials of improving bioconversion of lignocellulosic biomass like Miscanthus sinensis, hence making this crop a valuable energy plant.

Comparison of Corn Stover Pretreatments with Lewis Acid Catalyzed Choline Chloride, Glycerol and Choline Chloride-Glycerol Deep Eutectic Solvent.

Herein, corn stover (CS) was pretreated by less corrosive lewis acid FeCl3 acidified solutions of neat and aqueous deep eutectic solvent (DES), aqueous ChCl and glycerol at 120 °C for 4 h with single FeCl3 pretreatment as control. It was unexpected that acidified solutions of both ChCl and glycerol were found to be more efficient at removing lignin and xylan, leading to higher enzymatic digestibility of pretreated CS than acidified DES. Comparatively, acidified ChCl solution exhibited better pretreatment performance than acidified glycerol solution. In addition, 20 wt% water in DES dramatically reduced the capability of DES for delignification and xylan removal and subsequent enzymatic cellulose saccharification of pretreated CS. Correlation analysis showed that enzymatic saccharification of pretreated CS was highly correlated to delignification and cellulose crystallinity, but lowly correlated to xylan removal. Recyclability experiments of different acidified pretreatment solutions showed progressive decrease in the pretreatment performance with increasing recycling runs. After four cycles, the smallest decrease in enzymatic cellulose conversion (22.07%) was observed from acidified neat DES pretreatment, while the largest decrease (43.80%) was from acidified ChCl pretreatment. Those findings would provide useful information for biomass processing with ChCl, glycerol and ChCl-glycerol DES.

Lignocellulosic ethanol production from cotton stalk: an overview on pretreatment, saccharification and fermentation methods for improved bioconversion process

Cotton stalk is the most widely generated agricultural residue with lower economic importance, and can be employed as a feedstock in lignocellulosic biorefinery for the manufacture of bioethanol and other value-added bioproducts. Cotton stalk possesses high holocellulose content, which can be saccharified to various fermentable sugars for bioethanol production. However, the occurrence of high amount of lignin in cotton stalk renders it an inferior substrate for bioethanol production. Selection of suitable pretreatment process can improve digestibility of cotton stalk and hence higher sugar concentration on subsequent enzymatic saccharification. Furthermore, fermentation of hexose and pentoses sugars to ethanol requires robust microbial strains and efficient fermentation methods. Therefore, the major hindrance in commercializing lignocellulosic ethanol from cotton stalk is to develop an effective combination of pretreatment, saccharification, and fermentation methods thereby making the whole bioconversion process economically viable. This review paper discusses various previously investigated pretreatment, acid and/or enzymatic saccharification, and fermentation methods for cotton stalk-to-lignocellulosic ethanol production. Finally, it also discusses the major barriers in bioethanol fermentation strategies, and as well future perspectives to overcome these issues.

Co-production of fermentable glucose, xylose equivalents, and HBS-lignin from sugarcane bagasse through a FeCl3-catalyzed EG/H2O pretreatment

In this study, a FeCl3-catalyzed ethylene glycol/water (FeCl3-EG/H2O) pretreatment was developed and applied for the enzymatic hydrolysis of sugarcane bagasse. The results indicated that FeCl3-EG/H2O pretreatment showed excellent selectivity for the removal of both hemicellulose (∼100 %) and lignin (∼65.3 %), leading to the residual pretreated solid with high cellulose content (∼74.7 %) that could be easily for subsequent enzymatic saccharification. The yield of glucose after enzymatic hydrolysis was almost linearly positively correlated with the removal of lignin and xylan, and the maximum glucose yield obtained the using method was 91.5 %, which was 4.1-fold as large as that without pretreatment. The SEM, FT-IR, and XRD analysis indicated that the improvement of enzyme digestibility of sugarcane bagasse was not only related to the selectively remove of some component barriers, but was also correlated with the changes of surface morphology and internal structure. Furthermore, part of the hemicellulose and lignin in sugarcane bagasse also can be recovered in xylose equivalents, furfural, and high boiling solvent lignin (HBS-lignin) with the yields of 84.0 %, 12.5 %, and 60.1 %, respectively. It can be concluded that the FeCl3-EG/H2O pretreatment developed in this study was effective in achieving the hierarchic conversion and utilization of bagasse.

Hydrothermal Pretreatment of Wheat Straw: Effects of Temperature and Acidity on Byproduct Formation and Inhibition of Enzymatic Hydrolysis and Ethanolic Fermentation

Biochemical conversion of wheat straw was investigated using hydrothermal pretreatment, enzymatic saccharification, and microbial fermentation. Pretreatment conditions that were compared included autocatalyzed hydrothermal pretreatment at 160, 175, 190, and 205 °C and sulfuric-acid-catalyzed hydrothermal pretreatment at 160 and 190 °C. The effects of using different pretreatment conditions were investigated with regard to (i) chemical composition and enzymatic digestibility of pretreated solids, (ii) carbohydrate composition of pretreatment liquids, (iii) inhibitory byproducts in pretreatment liquids, (iv) furfural in condensates, and (v) fermentability using yeast. The methods used included two-step analytical acid hydrolysis combined with high-performance anion-exchange chromatography (HPAEC), HPLC, ultra-high performance liquid chromatography-electrospray ionization-triple quadrupole-mass spectrometry (UHPLC-ESI-QqQ-MS), and pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS). Lignin recoveries in the range of 108–119% for autocatalyzed hydrothermal pretreatment at 205 °C and sulfuric-acid-catalyzed hydrothermal pretreatment were attributed to pseudolignin formation. Xylose concentration in the pretreatment liquid increased with temperature up to 190 °C and then decreased. Enzymatic digestibility was correlated with the removal of hemicelluloses, which was almost quantitative for the autocatalyzed hydrothermal pretreatment at 205 °C. Except for the pretreatment liquid from the autocatalyzed hydrothermal pretreatment at 205 °C, the inhibitory effects on Saccharomyces cerevisiae yeast were low. The highest combined yield of glucose and xylose was achieved for autocatalyzed hydrothermal pretreatment at 190 °C and the subsequent enzymatic saccharification that resulted in approximately 480 kg/ton (dry weight) raw wheat straw.

Double bonus: surfactant-assisted biomass pelleting benefits both the pelleting process and subsequent enzymatic saccharification of the pretreated pellets

A novel biomass processing approach that adding surfactant during pelleting and using the pellets for pretreatment and enzymatic saccharification was proposed.

Storage and handling of pretreated lignocellulose affects the redox chemistry during subsequent enzymatic saccharification

The decomposition of lignocellulose in nature, as well as when used as feedstock in industrial settings, takes place in a dynamic system of biotic and abiotic reactions. In the present study, the impact of abiotic reactions during the storage of pretreated lignocellulose on the efficiency of subsequent saccharification was investigated. Abiotic decarboxylation was higher in steam-pretreated wheat straw (SWS, up till 1.5% CO2) than in dilute-acid-catalysed steam-pretreated forestry residue (SFR, up till 3.2% CO2) which could be due to higher iron content in SFR and there was no significant CO2 production in warm-water-washed slurries. Unwashed slurries rapidly consumed O2 during incubation at 50 °C; the behaviour was more dependent on storage conditions in case of SWS than SFR slurries. There was a pH drop in the slurries which did not correlate with acetic acid release. Storage of SWS under aerobic conditions led to oxidation of the substrate and reduced the extent of enzymatic saccharification by Cellic® CTec3. Catalase had no effect on the fractional conversion of the aerobically stored substrate, suggesting that the lower fractional conversion was due to reduced activity of the lytic polysaccharide monooxygenase component during saccharification. The fractional conversion of SFR was low in all cases, and cellulose hydrolysis ceased before the first sampling point. This was possibly due to excessive pretreatment of the forest residues. The conditions at which pretreated lignocellulose are stored after pretreatment significantly influenced the extent and kind of abiotic reactions that take place during storage. This in turn influenced the efficiency of subsequent saccharification. Pretreated substrates for laboratory testing must, therefore, be stored in a manner that minimizes abiotic oxidation to ensure that the properties of the substrate resemble those in an industrial setting, where pretreated lignocellulose is fed almost directly into the saccharification vessel.

A combined pretreatment with ultrasound-assisted alkaline solution and aqueous deep eutectic solvent for enhancing delignification and enzymatic hydrolysis from oil palm fronds

Oil palm frond (OPF) is one of the most abundant lignocellulosic biomass wastes found in the palm oil industry. However, the recalcitrant nature of OPF makes it hard to produce reducing sugar and convert it into subsequent high-value chemicals. Therefore, OPF was subjected to an ultrasound-assisted NaOH-aqueous deep eutectic solvent (DES) pretreatment in this study to increase delignification and enhance enzymatic saccharification. The effects of ultrasonication on the combined NaOH-aqueous DES for OPF pretreatment and saccharification were scrutinized. Under the recommended ultrasonic pretreatment (amplitude 60 %, duration 30 min) with 2.5 % NaOH-aqueous DES, lignin removal was significantly improved to 47.00 % in comparison to that obtained without ultrasound pretreatment, 30.92 %. OPF subjected to the recommended pretreatment was able to retain a high polysaccharide content, where subsequent enzymatic saccharification was able to achieve a glucose yield of 90.12 %. Physical and chemical characterizations of pretreated OPF were carried out to provide evidence to explain the observed improvement in lignin removal and enzymatic hydrolysis. The use of NaOH-aqueous DES as an ultrasonic medium was proven to be an effective method to promote a novel one-pot, two-processes pretreatment system.