Hydrolysis Yield Research Articles

Production of Medium Chain Length Polyhydroxyalkanoate from Waste Cannabis sativa Biomass

This works aims to evaluate a process to convert residual medical Cannabis sativa stalks into medium chain length polyhydroxyalkanoate (mcl-PHA), using chemical pretreatment and enzymatic hydrolysis to obtain sugars as a carbon source for a fermentation process with Pseudomonas aeruginosa. Chemical composition analysis revealed a content of structural polysaccharides of 57.64%. Thermochemical pretreatments with 2% sulfuric acid or 2% sodium hydroxide were capable of partially removing hemicellulose and lignin, as well as increasing cellulose crystallinity. Pretreated biomass was subjected to hydrolysis using commercial cellulase cocktails Celluclast® 1.5 L and Cellic® CTec3. Acid pretreatment showed an adverse effect on hydrolysis yield of holocellulose, decreasing to 39.5%; compared to 44.4% of untreated biomass. Alkaline pretreatment increased degree of hydrolysis up to 73.3%. Shake flask fermentation of hydrolysate with Pseudomonas aeruginosa produced cell growth of 1.65 g/L and a mcl-PHA titer of 0.41 g/L. Extracted polymer presented characteristic FTIR bands for PHAs, glass transition temperature of − 50.8 °C, melting temperature of 48.9 °C, possibly allowing its use in the biomedical industry. The developed process represents a potential way to valorize Cannabis waste stalks, using alkaline pretreatment, enzymatic hydrolysis with Celluclast® 1.5 L and fermentation with Pseudomonas aeruginosa. Further work should focus on improving yield of the obtained polyhydroxyalkanoate in fermentation, in order to improve industrial feasibility of the entire process and, in turn, increasing revenue of the medical Cannabis industry.Graphical

Acidogenic Fermentation of Food Waste for the Production of Short-Chain Fatty Acids: The Impact of Inoculum Type and Inoculum Heat Pretreatment

Acidogenic fermentation is an emerging biotechnology that allows for the utilization of food waste as a feedstock to produce high-value products such as short-chain fatty acids (SCFAs), effectively offering a tangible solution for food waste management as well as resource recovery. The objectives of the current study were to identify the ideal inoculum, waste-activated sludge (WAS) or anaerobic digester sludge (AD), for the acidogenic fermentation of food waste at room temperature, as well as to evaluate the impact of heat pretreatment of these inoculums on fermentation performance. The maximum hydrolysis yield of 399 g sCOD/kg VS added was obtained when untreated AD was used as the inoculum, whereas the pretreated AD inoculum provided the highest SCFA yield and conversion efficiency of 238 g sCODSCFA/kg VS added and 71%, respectively. Heat pretreatment had a detrimental impact on the WAS inoculum, leading to lower hydrolysis and SCFA yields, but exerted a positive influence on the AD inoculum. The microbial community showed that heat pretreatment negatively impacted the abundance of non-spore-forming hydrolytic and acidogenic microorganisms. Overall, this study demonstrates the critical role of inoculum type and heat pretreatment in optimizing the acidogenic fermentation process, laying the groundwork for future refinements in SCFA production from food waste through inoculum design.

Protein hydrolysates prepared by Alcalase using ultrasound and microwave pretreated almond meal and their characterization.

The study aimed to optimize ultrasonic (US: 40kHz/200W for 10, 20, 30, 40, and 50min), and microwave (MW: 160W for 45, 90, 125, 180, and 225s) pretreatment conditions on protein extraction yield and degree of protein hydrolysis (DH) from almond de-oiled meal, an industrial by-product. First order model was used to describe the kinetics of almond protein hydrolysates obtained with Alcalase. The highest DH, 10.95% was recorded for the US-50min and 8.87% for MW-45s; while it was 5.76% for the untreated/control sample. At these optimized pretreatment conditions, a 1.16- and 1.18-fold increment in protein recovery was observed for theUS and MW pretreatments, respectively in comparison to the conventional alkaline extraction. The molecular weight distribution recorded for pretreated samples disclosed a significant reduction in the band thickness in comparison with control. Both the pretreatments resulted in a significant increase (P < 0.05) in the antioxidant activity, and TCA solubility index when compared with the control. Results evinced that US and/or MW pretreatments before enzymatic hydrolysis can be a promising approach for the valorization of almond meal for its subsequent useas an ingredient for functional foods/nutraceuticals which otherwise fetches low value as an animal feed.

Hydrolysis behavior and mechanistic insights of CaMg2InX (X = 0.1, 0.3, 0.5, 0.7) ternary alloy for sustainable hydrogen production

The hydrolysis behavior of CaMg2In0.1, CaMg2In0.3, CaMg2In0.5, and CaMg2In0.7 ternary alloys in an MgCl2 solution following casting and hydrogenation were investigated. The hydrolysis mechanism of these alloys is elucidated through an analysis of microstructure, phase composition, and kinetics before and after hydrolysis. The nucleation-growth Avrami model is employed to accurately model the hydrolysis kinetics, revealing improved hydrolysis yields and reaction rates following hydrogenation. Notably, CaMg2In0.1 has demonstrated exceptional hydrolysis characteristics, exhibiting a yield of 1140 mL/g, an initial hydrolysis rate of 113 mL/g·s, and an activation energy of 24.3 ± 1.7 kJ·mol−1. The yield of H-CaMg2In0.1 further escalates to 1800 mL/g with a rate of 221 mL/g·s, attributed to the formation of Ca4Mg3H14 and In phases subsequent to the hydrogenation of In2Ca and Mg3In phases in the alloy. These newly formed phases act as catalysts and actively participate in the hydrolysis process, providing active sites for hydrogen production, thus enhancing hydrolysis yields and kinetics. It is observed that with increasing In content, the order of hydrolysis performance of the alloy is as follows: CaMg2In0.1 > CaMg2In0.3 > CaMg2In0.5 > CaMg2In0.7, consistent with the trend after hydrogenation. These findings indicate that the addition of In significantly enhances the hydrolysis performance of CaMg2 alloys, offering a promising strategy for preparing magnesium-based alloys with high yields and favorable kinetic properties.

Encouraging Solution to the Problem of Synthesizing Protein α‐Thioester

Comprehensive SummaryExpressed protein ligation (EPL) provides a powerful tool to access large‐size proteins with precise structures. Existing methods for constructing the critical protein thioester for EPL have predominantly relied on the recombinant intein fusion expressed in Escherichia coli (E. coli). Despite its powerful applications, the expression of thioester derived from eukaryotic protein in E. coli inherently suffers from its limited solubility, the inactivity of intein, premature hydrolysis and low yields. To overcome these obstacles, we present herein the facile one‐flask synthesis of inaccessible protein α‐thioester via a SUMO‐protein‐intein (SPI) sandwich model. The utility of SUMO enhances the protein fusion yield and solubility, prevents premature hydrolysis and simplifies the purification process. The inaccessible protein thioester with internal Cys residues can be readily produced and is compatible with the EPL‐desulfurization protocol used to prepare complex proteins, which is otherwise difficult to obtain using traditional methods. Its utility has been highlighted through the synthesis of human granulocyte colony‐stimulating factor (G‐CSF).

Comparative study for enzymatic hydrolysis of sugarcane bagasse using free and nanoparticle immobilized holocellulolytic enzyme cocktail

This study aimed at a comparative analysis of enzymatic hydrolysis of sugarcane bagasse biomass by using a free holocellulase cocktail of an enzyme (Cellic CTec2) and immobilized enzyme system on IOMNP@SiO2-NH2 support to make the process cost-effective. The immobilization was substantiated by examining the nano biocatalysts using FTIR, TEM, and DLS analysis. IOMNP@SiO2 –NH2 and immobilized enzyme system have average particle sizes of 81.1, and 455.9 nm confirmed by DLS. Response surface methodology (RSM) based on the Box Behnken design was employed to optimize the saccharification process conditions. The free cellulase cocktail showed a maximum hydrolysis yield of 70.71 % at 12.5FPU/gm enzyme dose, while the immobilized enzyme at a dose of 20 FPU/gm achieved a conversion rate of 74.19 % under optimized parameters likes hydrolysis time, surfactant and substrate concentration. Pretreated sugarcane bagasse substrate was used to evaluate the reusability potential of magnetically immobilized cellulase enzyme compared to recovered free enzyme up to 5 cycles. In the second and third cycles, the conversion rates of cellulose to glucose were determined to be 68.21 %, and 52.6 %, respectively. The findings of this study demonstrate that the immobilized Cellic CTec2 enzyme cocktail on functionalized IOMNP is effective for hydrolyzing sugarcane bagasse which also provides the advantages of easy recovery and reusability of enzyme for multiple hydrolysis cycles.

Efficient Fast Fractionation of Biomass Using a Diol-Based Deep Eutectic Solvent for Facilitating Enzymatic Hydrolysis and Obtaining High-Quality Lignin.

Current DES pretreatment is often performed under relatively severe conditions with high temperature, long time, and high DES usage. This work studied a short-time diol DES (deep eutectic solvent) pretreatment under mild conditions to fractionate the bamboo, facilitate enzymatic hydrolysis, and obtain high-quality lignin. At an optimized condition of 130 °C for only 10 min, lignin and xylan removal reached 61.34 % and 84.15 %, with residual glucan showing a ~90 % enzymatic hydrolysis yield. Equally important, the dissolved lignin could be readily recovered with 97.51 % yield, exhibiting 96.65 % β-O-4 preservation. The fractionation and lignin protection mechanisms were unveiled by XRD, FTIR, cellulose-DP, 2D HSQC NMR, 31P NMR and GPC analysis. This study highlighted that short-time fractionation of bamboo can be achieved by a diol-based DES which is an ideal strategy to upgrade the lignocellulose biomass for high enzymatic hydrolysis yields and high-quality lignin stream.

Optimizing bioconversion processes of rice husk into value-added products: D-psicose, bioethanol, and lactic acid

Rice husk, rich carbon content, is an agricultural waste produced globally at an amount of 120 million tons annually, and it has high potential as a biorefinery feedstock. Herein, we investigated the feasibility of producing various products as D-psicose, bioethanol and lactic acid from rice husk (RH) through a biorefinery process. Alkali-hydrogen peroxide-acetic acid pretreatment of RH effectively removed lignin and silica, resulting in enzymatic hydrolysis yield of approximately 86.3% under optimal hydrolysis conditions. By using xylose isomerase as well as D-psicose-3-epimerase with borate, glucose present in the RH hydrolysate was converted into D-psicose with a 40.6% conversion yield in the presence of borate. Furthermore, bioethanol (85.4%) and lactic acid (92.5%) were successfully produced from the RH hydrolysate. This study confirmed the high potential of RH as a biorefinery feedstock, and it is expected that various platform chemicals and value-added products can be produced using RH.

Structural characterization of lignin from the green pretreatments for co-producing xylo-oligosaccharides and glucose: Toward full biomass utilization

Three green non-enzymatic catalysis pretreatments (NECPs) including autohydrolysis, subcritical CO2-assisted seawater autohydrolysis, and inorganic salt catalysis were utilized to simultaneously produce xylo-oligosaccharides (XOS), glucose, and cellulolytic enzyme lignin (CEL) from sugarcane bagasse (SCB). The yield of XOS in all three NECPs was over 50 % with a competitive glucose yield of enzymatic hydrolysis. And the effects of different pretreatments on the chemical structure and composition of CEL samples were also investigated. The pretreatments significantly increased the thermal stability, yield, and purity of the CEL samples. Moreover, the net yield of lignin was 58.3 % with lignin purity was 98.9 % in the autohydrolysis system. Furthermore, there was a decrease in the molecular weight of CEL samples as the pretreatment intensity increased. And the original lignin structural units sustained less damage during the NECPs, due to the cleavage of the β-O-4 bonds dominating lignin degradation. Meanwhile, these pretreatments increased the phenolic-OH in CEL samples, making the lignin more reactive, and enhancing its subsequent modification and utilization. Collectively, the described techniques have demonstrated practical significance for the coproduction of XOS and glucose, and lignin, providing a promising strategy for full utilization of biomass.

Estimation of applicability of protein hydrolysate of black soldier fly (Hermetia illucens) larvae prepared using enzymatic hydrolysis as a fish feed component

The special advantage of black soldier fly (BSF) (Hermetia illucens) larvae for use as feed is their high protein content and low chitin content, which makes the larvae easier to digest, making them nutritious for fish, farm animals and birds. It has been reported that BSF larvae contain all the amino acids necessary for animal protein, including the essential amino acids, also macro- and micronutrients. The technological parameters of BSF larvae protein hydrolysis procedure were optimized. Some physicochemical parameters of the hydrolysate and the yield of hydrolysis were estimated for the hydrolysate prepared according to the optimized parameters. The sterlet (Acipenser ruthenus) starter feed having 10% of its fishmeal portion substituted with BSF larvae protein hydrolysate has been tested. The BSF larvae protein hydrolysate may be included in fish feed formulations, especially in starter feeds.

Gamma radiation vs high pressure pretreatment on physicochemical characteristics of rice bran hydrolysate

This study investigated the effect of using gamma radiation and high-pressure processing as pretreatment, to consider the structural and amino acid composition changes in rice bran hydrolysate (RBH). The extraction yield and degree of hydrolysis of the irradiated sample were greater than those of the pressurized and control samples, which radiation at 10 kGy gave 31 % yield. Protein content of the control was the highest at 36.1 %, with 32.4 % in pressurized sample at 500 MPa. Control had the highest concentration of total and branched-chain amino acids, with a value of 25,834 mg/100g. Before and after extraction, the microstructure changed visibly and protein agglomeration can be significantly induced by applying a high-pressure. Therefore, this study showed the potential of using both pretreatment methods prior to enzymolysis extraction, with radiation producing more extract. High-pressure produced more protein content, but neither method produced any difference in amino acid content.

Effect of various aromatic compounds with different functional groups on enzymatic hydrolysis of microcrystalline cellulose and alkaline pretreated wheat straw

Low molecular aromatic compounds are detrimental to the enzymatic hydrolysis of lignocellulose. However, the specific role of their functional groups remains unclear. Here, a series of nine aromatic compounds as additives were tested to understand their effect on the hydrolysis yield of microcrystalline cellulose (MCC) and alkaline pretreated wheat straw. Based on the results, the inhibition of aldehyde groups on MCC was greater than that of carboxyl groups, whereas for the alkaline pretreated wheat straw case, the inhibitory effect of aldehyde groups was lower than that of carboxyl groups. Increased methoxyl groups of aromatic compounds reduced the inhibitory effect on enzymatic hydrolysis of both substrates. Stronger inhibition of aromatic compounds on MCC hydrolysis was detected in comparison with the alkaline pretreated wheat straw, indicating that the substrate lignin can offset the inhibition to a certain extent. Among all aromatic compounds, syringaldehyde with one aldehyde group and two methoxyl groups improved the glucan conversion of the alkaline pretreated wheat straw.

Hydrodynamic cavitation-assisted acid pretreatment and fed-batch simultaneous saccharification and co-fermentation for ethanol production from sugarcane bagasse using immobilized cells of Scheffersomyces parashehatae

A new alternative for hydrodynamic cavitation-assisted pretreatment of sugarcane bagasse was proposed, along with a simultaneous saccharification and co-fermentation (SSCF) process performed in interconnected columns. Influential variables in the pretreatment were evaluated using a statistical design, indicating that an ozone flow rate of 10 mg min−1 and a pH of 5.10 resulted in 86 % and 72 % glucan and xylan hydrolysis yields, respectively, in the subsequent enzymatic hydrolysis process. Under these optimized conditions, iron sulfate (15 mg L−1) was added to assess Fenton pretreatment, resulting in glucan and xylan hydrolysis yields of 92 % and 71 %, respectively, in a material pretreated for 10 min. In SSCF, ethanol volumetric productivities of 0.33 g L−1 h−1 and of 0.54 g L−1 h−1 were obtained in batch and fed-batch operation modes, achieving 26 g L−1 of ethanol in 48 h in the latter mode.

In Vitro Stability of ACE-Inhibitory Peptides of Rohu Fish Waste to Heat, pH and Gastrointestinal Enzymes

Angiotensin-I converting enzyme inhibitory (ACEi) peptides were prepared from rohu (Labeo rohita) fish wastes by enzymatic hydrolysis [1.1% alcalase concentration (v/w); 130 min time; 52°C temperature and 0.8:1 solid-liquid ratio]. Ultrasound Assisted Enzymatic Extraction (UAEE) and Microwave Assisted Enzymatic Extraction (MAEE) were performed to enhance degree of hydrolysis (DH), ACEi activity, and peptide yield (PY). The ACEi peptides were examined for stability during thermal processing, varied pH treatments and in vitro gastrointestinal digestion. Peptides were stable at different temperatures (25, 37, 55, 75, and 100°C) except at 121°C which reduced their stability by 7%, 6.9% and 2.2% in UAEE, MAEE and Non-Assisted Enzymatic Extraction (NAEE) peptides, respectively. Similarly, peptides retained ACEi at pH 2 to 8 and subsequent reduction at pH&gt;8. In vitro digestion studies showed stability of all peptides to GI digestion at 0.05% pepsin, 0.05% trypsin, 0.05% chymotrypsin and the combination of 0.025% trypsin and 0.025% chymotrypsin except MAEE peptides showing less stability with pepsin (0.05%). Therefore, it is concluded that these ACEi peptides derived from rohu fish waste, stable to heat, pH and gastrointestinal enzymatic action, can be used as ingredient in functional foods, which undergo thermal processing and digestion.

Cloning and expression of Neurospora crassa cellobiohydrolase II in Pichia pastoris

A major cellobiohydrolase of Neurospora crassa CBH2 was successfully expressed in Pichia pastoris. The maximum Avicelase activity in shake flask among seven transformants which selected on 4.0 g/L G418 plates was 0.61 U/mL. The optimal pH and temperature for Avicelase activity of the recombinant CBH2 were determined to be 4.8 and 60 °C, respectively. The new CBH2 maintained 63.5 % Avicelase activity in the range of pH 4.0–10.4, and 60.2 % Avicelase activity in the range of 30–90 °C. After incubation at 70–90 °C for 1 h, the Avicelase activity retained 60.5 % of its initial activity. The presence of Zn2+, Ca2+ or Cd2+ enhanced the Avicelase activity of the CBH2, of which Cd2+ at 10 mM causing the highest increase. The recombinant CBH2 was used to enhance the Avicel hydrolysis by improving the exo-exo-synergism between CBH2 and CBH1 in N.crassa cellulase. The enzymatic hydrolysis yield was increased by 38.1 % by adding recombinant CBH2 and CBH1, and the yield was increased by 215.4 % when the temperature is raised to 70 °C. This work provided a CBH2 with broader pH range and better heat resistance, which is a potential enzyme candidate in food, textile, pulp and paper industries, and other industrial fields.

Physicochemical characterization of açaí seeds (Euterpe oleracea) from Colombian pacific and their potential of mannan-oligosaccharides and sugar production via enzymatic hydrolysis

AbstractThe study aimed to characterize açaí seeds and explore their potential for producing mannooligosaccharides (MOS) through enzymatic hydrolysis. According to characterization tests by XRD, FTIR, and chemical analysis, acai seeds, a waste material from acai fruit processing, contain significant hemicellulose content, with a main content based on mannan. The study utilized Rohalase®GMP enzyme for hydrolysis and monitored reducing sugars and MOS production (8–10 g MOS/L) hydrolysate with varying lengths, including di-, tri-, tetra-, and pentasaccharide over time. Results showed a concentration plateau of reducing sugars at 13 h and increased MOS until hour nineteen. Mannobiose (M2) was the predominant MOS produced. Comparative analysis with prior research indicated that the açaí seed hydrolysate’s MOS content aligns with that from other sources. Importantly, the study achieved % hydrolysis and MOS yield of 10.79 ± 0.05% and 51.39 ± 0.11 g M2-M5/g mannan, respectively. Açaí seed is a promising source to produce enzymes, biofuels, or thermal energy, as well as highly valued chemical compounds in the industry. This work additionally demonstrates its potential as a sustainable source of valuable MOS, suggesting applications as prebiotics and functional food additives, with implications for various industries seeking eco-friendly alternatives.

Enhancement of Liquid Hot Water Pretreatment on Corn Stover with Ball Milling to Improve Total Sugar Yields

Conversion of the lignocellulosic biomass to bioethanol contributes to the reduction of greenhouse gas emissions, enhancement of energy security, utilization of waste materials, and the promotion of sustainable agricultural practices. In this study, we report the effect of combining ball milling followed by liquid hot water (LHW) pretreatment of corn stover to lower the amount of enzyme required while also greatly increasing the recovery of xylose in fermentable form compared to either pretreatment alone. Short-duration ball milling for 60 min reduces the particle size of corn stover to 37.3 μm; however, the glucose only increased to 47% compared to 32% for unpretreated corn stover. In contrast, liquid hot water pretreatment alone can achieve increasing enzyme hydrolysis yields of cellulose from 49% to 93% as the pretreatment severity factor is increased from 3.24 to 4.41. However, the xylose yield decreased to 36% due to the fact that a considerable part of the xylose was degraded into furfural and humins. Surprisingly, the combination of mild ball milling (30 min) followed by mild liquid hot water pretreatment (190 °C, 15 min) could achieve both high glucose (83%) and xylose (72%) yields for a total sugar yield of 79%, theoretically. Thus, combining ball milling with liquid hot water pretreatment allows for milder conditions for both processes that lead to enhanced cellulose conversion without sacrificing xylose to degradation, which hinders enzymatic hydrolysis.

Alleviating lignin repolymerization facilitated the combined acid and deep eutectic solvent pretreatment of Populus

To elucidate the effects of lignin repolymerization on the combined pretreatment, the sequential acid and deep eutectic solvent (DES) pretreatment was implemented on Populus (poplar) in this study. Effects of acid pretreatment with or without phenolic additives (2-naphthol (2 N) and resorcinol (RS)) on the DES post-treatment efficiency were examined. The results showed that 2 N inhibited the lignin repolymerization in the first-step pretreatment, and slightly enhanced the lignin removal during DES post-treatment. However, RS intensified the lignin repolymerization, significantly decreasing the delignification of DES post-treatment from 73.6%∼79.4% to 39.0∼43.0%. Moreover, the poorer enzymatic hydrolysis was observed in the combined pretreated substrates as compared to the single DES pretreated ones, due to the higher surface lignin coverage. The addition of 2 N counteracted the adverse effects of acid pretreatment on the hydrolysis yields of combined pretreated substrates, and facilitated the yields from 25.8%∼36.0% to 26.2∼67.3%, while the RS addition further decreased the hydrolysis yields to 8.6∼9.6%. The mechanism for the changes on enzymatic digestibilities of the combined pretreated substrates was elucidated.

Fermentative Production of β-Carotene from Sugarcane Bagasse Hydrolysate by Rhodotorula glutinis CCT-2186.

Β-Carotene is a red-orange pigment that serves as a precursor to important pharmaceutical molecules like vitamin A and retinol, making it highly significant in the industrial sector. Consequently, there is an ongoing quest for more sustainable production methods. In this study, glucose and xylose, two primary sugars derived from sugarcane bagasse (SCB), were utilized as substrates for β-carotene production by Rhodotorula glutinis CCT-2186. To achieve this, SCB underwent pretreatment using NaOH, involved different concentrations of total solids (TS) (10%, 15%, and 20%) to remove lignin. Each sample was enzymatically hydrolyzed using two substrate loadings (5% and 10%). The pretreated SCB with 10%, 15%, and 20% TS exhibited glucose hydrolysis yields (%wt) of 93.10%, 91.88%, and 90.77%, respectively. The resulting hydrolysate was employed for β-carotene production under batch fermentation. After 72h of fermentation, the SCB hydrolysate yielded a β-carotene concentration of 118.56 ± 3.01mg/L. These findings showcase the robustness of R. glutinis as a biocatalyst for converting SCB into β-carotene.

Stepwise transform sugarcane bagasse into xylooligosaccharides and fermentable glucose by hydrothermal-xylanase-acid-cellulase hydrolysis

Hydrothermal pretreatment can effectively degrade the hemicellulosic xylan into xylooligosaccharides; however, the low degree of polymerization (DP) oligomers are dissatisfied. Considering the significance of low DP, one approach to enhance the levels of low DP xylooligosaccharides is to utilize endo-xylanase for degrading the high DP xylooligosaccharides. Therefore, a combination method of hydrothermal and post endo-xylanase hydrolysis was employed in this study to enhance XOS yields. As a result, a maximum XOS of 54.5% with low DP was achieved from sugarcane bagasse, and the ratio of (DP2+DP3)/XOS was remarkably improved. Following the preparation of XOS, an additional acidic hydrolysis using dilute sulfuric acid was performed to enhance the saccharification efficiency of cellulose, resulting in a significant increase in yield of enzymatic hydrolysis to over 90%. Overall, biomass-derived sugar products (XOS and glucose) can be co-produced for adding the value of the sugarcane bagasse by sequential hydrothermal-endo-xylanase-acid-cellulase hydrolysis.