Hailong Li

Rapid analysis of mono-saccharides and oligo-saccharides in hydrolysates of lignocellulosic biomass by HPLC

HPLC using pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) was used to analyse mono-saccharides and oligo-saccharides in hydrolysates of lignocellulosic biomass. PMP derivatives, including those of mannose, rhamnose, cellobiose, glucose, xylose and arabinose, were separated within 14xa0min with detection at 254xa0nm. The method was also suitable for xylo-oligosaccharides (XOS): PMP derivatives of xylohexaose, xylopentaose, xylotetraose, xylotriose and xylobiose were well separated under the same conditions. The method was used to determine the mono-saccharide composition of Miscanthus and evaluate the production of XOS from enzymatic hydrolysis of crude xylan.

Adsorption behavior of levulinic acid onto microporous hyper-cross-linked polymers in aqueous solution: Equilibrium, thermodynamic, kinetic simulation and fixed-bed column studies

The recovery of levulinic acid (LA) from aqueous solution and actual biomass hydrolysate by a microporous hyper-cross-linked polymer, SY-01, was investigated for the first time under batch and fixed-bed column conditions. The results showed that the optimum pH should be in the acidic range (pHxa0<xa03.0) without adjusting the pH. In the single-component system equilibrium study, the Langmuir isotherm model fits the LA adsorption onto SY-01 resin better than the Freundlich isotherm model, indicating that LA adsorption onto SY-01 resin under the concentration range studied is a monolayer homogeneous adsorption process. The maximum adsorption capacity of LA onto SY-01 resin decreased with increasing temperature, ranging from 103.74 to 95.70xa0mg/g. The obtained thermodynamic parameters suggested that the adsorption of LA on SY-01 was spontaneous (ΔG0<-3.788xa0kJ/mol), and exothermic (ΔH0xa0=xa0-11.764xa0kJ/mol). For kinetic study, the adsorption of LA onto SY-01 resin at various operating conditions follows the pore diffusion model and the intraparticle diffusion is the rate-limiting step for the adsorption of LA onto SY-01 resin. The effective pore diffusivity was dependent upon temperature, but independent of initial LA concentration, and were 3.306xa0×xa010-10, 5.274xa0×xa010-10 and 7.707xa0×xa010-10xa0m2/s at 298, 318 and 338xa0K, respectively. In desorption process, the recovery efficiency of LA from SY-01 resin was 99.39%, and LA concentration in the eluent was raised 2.97-fold. In conclusion, our results show that the SY-01 resin has potential application in product recovery of LA from biomass hydrolysate.

Enhanced enzymatic hydrolysis and acetone-butanol-ethanol fermentation of sugarcane bagasse by combined diluted acid with oxidate ammonolysis pretreatment.

This study aims to propose a biorefinery pretreatment technology for the bioconversion of sugarcane bagasse (SB) into biofuels and N-fertilizers. Performance of diluted acid (DA), aqueous ammonia (AA), oxidate ammonolysis (OA) and the combined DA with AA or OA were compared in SB pretreatment by enzymatic hydrolysis, structural characterization and acetone-butanol-ethanol (ABE) fermentation. Results indicated that DA-OA pretreatment improves the digestibility of SB by sufficiently hydrolyzing hemicellulose into fermentable monosaccharides and oxidating lignin into soluble N-fertilizer with high nitrogen content (11.25%) and low C/N ratio (3.39). The enzymatic hydrolysates from DA-OA pretreated SB mainly composed of glucose was more suitable for the production of ABE solvents than the enzymatic hydrolysates from OA pretreated SB containing high ratio of xylose. The fermentation of enzymatic hydrolysates from DA-OA pretreated SB produced 12.12g/L ABE in 120h. These results suggested that SB could be utilized efficient, economic, and environmental by DA-OA pretreatment.

CaCO3 supplementation alleviates the inhibition of formic acid on acetone/butanol/ethanol fermentation by Clostridium acetobutylicum

ObjectiveTo investigate the inhibiting effect of formic acid on acetone/butanol/ethanol (ABE) fermentation and explain the mechanism of the alleviation in the inhibiting effect under CaCO3 supplementation condition.ResultsFrom the medium containing 50xa0g sugars l−1 and 0.5xa0g formic acid l−1, only 0.75xa0g ABE l−1 was produced when pH was adjusted by KOH and fermentation ended prematurely before the transformation from acidogenesis to solventogenesis. In contrast, 11.4xa0g ABE l−1 was produced when pH was adjusted by 4xa0g CaCO3 l−1. The beneficial effect can be ascribed to the buffering capacity of CaCO3. Comparative analysis results showed that the undissociated formic acid concentration and acid production coupled with ATP and NADH was affected by the pH buffering capacity of CaCO3. Four millimole undissociated formic acid was the threshold at which the transformation to solventogenesis occurred.ConclusionThe inhibiting effect of formic acid on ABE fermentation can be alleviated by CaCO3 supplementation due to its buffering capacity.

Heterogeneous expression and functional characterization of cellulose-degrading enzymes from Aspergillus niger for enzymatic hydrolysis of alkali pretreated bamboo biomass.

AbstractEnzymatic hydrolysis of cellulosic biomass has caught much attention because of modest reaction conditions and environment friendly conditions. To reduce the cost and to achieve good quantity of cellulases, a heterologous expression system is highly favored. nIn this study, cellulose-degrading enzymes, GH3 family β-glucosidase (BGL), GH7 family-related cellobiohydrolases (CBHs), and endoglucanase (EG) from a newly isolated Aspergillus niger BE-2 are highly expressed in Pichia pastoris GS115. The strain produced EG, CBHs, and BGL enzymatic concentration of 0.56, 0.11, and 22xa0IU/mL, respectively. Mode of actions of the recombinant enzymes for substrate specificity and end product analysis are verified and found specific for cellulose degradation. Bamboo biomass saccharification with A. niger cellulase released a high level of fermentable sugars. Hydrolysis parameters are optimized to obtain reducing sugars level of 3.18xa0g/L. To obtain reducing sugars from a cellulosic biomass, A. niger could be a good candidate for enzymes resource of cellulase to produce reducing sugars from a cellulosic biomass. This study also facilitates the development of highly efficient enzyme cocktails for the bioconversion of lignocellulosic biomass into monosaccharides and oligosaccharides.

Extraction and characterization of wax from sugarcane bagasse and the enzymatic hydrolysis of dewaxed sugarcane bagasse

ABSTRACT Extraction of high-value products from agricultural wastes is an important component for sustainable bioeconomy development. In this study, wax extraction from sugarcane bagasse was performed and the beneficial effect of dewaxing pretreatment on the enzymatic hydrolysis was investigated. About 1.2% (w/w) of crude sugarcane wax was obtained from the sugarcane bagasse using the mixture of petroleum ether and ethanol (mass ratio of 1:1) as the extraction agent. Results of Fourier-transform infrared characterization and gas chromatography–mass spectrometry qualitative analysis showed that the crude sugarcane wax consisted of fatty fractions (fatty acids, fatty aldehydes, hydrocarbons, and esters) and small amount of lignin derivatives. In addition, the effect of dewaxing pretreatment on the enzymatic hydrolysis of sugarcane bagasse was also investigated. The digestibilities of cellulose and xylan in dewaxed sugarcane bagasse were 18.7 and 10.3%, respectively, compared with those of 13.1 and 8.9% obtained from native sugarcane bagasse. The dewaxed sugarcane bagasse became more accessible to enzyme due to the disruption of the outermost layer of the waxy materials.

The hydrolytic efficiency and synergistic action of recombinant xylan-degrading enzymes on xylan isolated from sugarcane bagasse

Understanding the interaction mechanisms between xylan and xylan-degrading enzymes is beneficial to the efficient hydrolysis of xylan. Xylan from sugarcane bagasse (SB) was extracted and characterized. The effects of heat treatment and removal of side chains of SB xylan on the hydrolytic efficiency and synergistic action of endo-β-1,4-xylanases (HoXyn11A and AnXyn10C), β-xylosidases (AnXln3D), and α-l-arabinofuranosidases (AnAxh62A) were investigated. Results indicated that heat treatment of xylan can improve the hydrolytic efficiency of xylan-degrading enzymes, and it is essential for the efficient hydrolysis of xylan by HoXyn11A. The removal of arabinofuranosyl side chains of xylan by AnAxh62A before enzymatic hydrolysis reduced the hydrolytic efficiency of HoXyn11A and AnXyn10C on xylan. AnXyn10C was more efficient than HoXyn11A in hydrolysis of xylan, whereas HoXyn11A showed better synergistic action than AnXyn10C with AnAxh62A and AnXln3D in the hydrolysis of xylan. This study provides new insights on the enzymatic hydrolysis of SB into monosaccharides and xylo-oligosaccharides.

Heterologous Expression and Characterization of an Acidic GH11 Family Xylanase from Hypocrea orientalis

A gene encoding glycoside hydrolase family 11 xylanase (HoXyn11B) from Hypocrea orientalis EU7–22 was expressed in Pichia pastoris with a high activity (413xa0IU/ml). HoXyn11B was partly N-glycosylated and appeared two protein bands (19–29xa0kDa) on SDS-PAGE. The recombinant enzyme exhibited optimal activity at pH 4.5 and 55xa0°C, and retained more than 90% of the original activity after incubation at 50xa0°C for 60xa0min. The determined apparent Km and Vmax values using beechwood xylan were 10.43xa0mg/ml and 3246.75xa0IU/mg, respectively. The modes of action of recombinant HoXyn11B on xylo-oligosaccharides (XOSs) and beechwood xylan were investigated by thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), which indicated that the modes of action of HoXyn11B are different from HoXyn11A since it is able to release a significant amount of xylose from various substrates. This study provides an opportunity to better understand the hydrolysis mechanisms of xylan by xylanases from Trichoderma.

Efficient Using Durian Shell Hydrolysate as Low-Cost Substrate for Bacterial Cellulose Production by Gluconacetobacter xylinus

Abstract Durian is one important tropical fruit with high nutritional value, but its shell is usually useless and considered as waste. To explore the efficient and high-value utilization of this agricultural and food waste, in this study, durian shell was simply hydrolyzed by dilute sulfuric acid, and the durian shell hydrolysate after detoxification was used for bacterial cellulose (BC) production by Gluconacetobacter xylinus for the first time. BC was synthesized in static culture for 10xa0days and the highest BC yield (2.67xa0g/L) was obtained at the 8th day. The typical carbon sources in the substrate including glucose, xylose, formic acid, acetic acid, etc. can be utilized by G. xylinus. The highest chemical oxygen demand (COD) removal (16.40%) was obtained at the 8th day. The highest BC yield on COD consumption and the highest BC yield on sugar consumption were 93.51% and 22.98% (w/w), respectively, suggesting this is one efficient bioconversion for BC production. Durian shell hydrolysate showed small influence on the BC structure by comparison with the structure of BC generated in traditional Hestrin–Schramm medium detected by FE-SEM, FTIR, and XRD. Overall, this technology can both solve the issue of waste durian shell and produce valuable bio-polymer (BC).

Improvement and Characterization in Enzymatic Hydrolysis of Regenerated Wheat Straw Dissolved by LiCl/DMAc Solvent System

Lithium chloride (LiCl)/N,N-dimethylacetamide (DMAc) solvent system was used to dissolve native and pretreated wheat straw materials in order to promote the enzymatic hydrolysis process. The dissolution ratio of wheat straw in LiCl/DMAc solvent system increased when dilute sulfuric acid or ethanol-sulfuric acid mixture pretreatment was conducted before dissolution. The materials regenerated from LiCl/DMAc solution exhibited obvious changes in structure and morphology, as revealed by scanning electron microscopy (SEM) and X-ray diffraction (XRD) analysis. The cellulose and xylan digestibilities of regenerated materials were improved obviously. The cellulose digestibilities of materials regenerated from native and pretreated wheat straw by dilute sulfuric acid and ethanol-sulfuric acid mixture were 61.8, 81.2, and 84.4xa0%, respectively, with 25 FPU of cellulase and 187xa0IU of xylanase after 96xa0h. These values were significantly higher than the digestibilities of 0, 33.2, and 57.5xa0% obtained from corresponding materials before dissolution. Dissolution treatment of wheat straw by LiCl/DMAc solvent system provides an alternative method for efficient enzymatic hydrolysis.