Structure-function relationship of licorice (Glycyrrhiza glabra) root extract-xanthan/guar gum mixture in a high sugar content system.

BACKGROUND: Foam-gels are one of the most important multicomponent-model systems in aerated confectionery, and an investigation of their microstructure is desirable. In this research, the structure-function relationship of xanthan gum/guar gum (XG/GG) and licorice (Glycyrrhiza glabra) root extract powder (LEP) was investigated in a high-sugar medium. Foam-gel systems were prepared at 4:10% to 8:20% ratios of LEP to biopolymer. RESULTS: The results show that increasing the LEP content reduced both the melting point and enthalpy, probably due to higher overrun and weaker junctions. Boosting the XG/GG ratio led the enhancement of mechanical properties, whereas increasing the LEP concentration weakened all textural parameters, which could be due to the poor structure of the network in the presence of the foaming agent, increased moisture content and overrun. In the whipped mixture samples containing 10 g kg-1 XG/GG, higher foaming capacity was observed. By increasing the level of biopolymers, smaller and more uniform air cells were formed according to a scanning electron microscopical study. At higher concentration of LEP, smaller bubbles and increased porosity were seen, which could be attributed to the availability of surfactant in the interfacial layer. CONCLUSION: Maximum structural strength was achieved at a 4:20 ratio of LEP to XG/GG. In rheological experiments, pseudoplastic behavior was seen in all samples. Generally, this model system can be simulated for other herbal extracts containing natural surfactants such as saponins. Achieving a more detailed understanding of these structures and their interactions could help in formulating novel food products. © 2021 Society of Chemical Industry.

Nitrate and nitrite pathways and dynamic changes in bacterial communities during beet sugar processing.

BACKGROUND: Bacterial community successions were surveyed during the processing stages of sugar production using high-throughput sequencing methods. Furthermore, the correlation between bacterial community and nitrate/nitrite content in beet sugar processing were investigated. RESULTS: In an analysis of the V3-V4 region of the 16S rDNA gene, 254 122 effective sequences were obtained from samples, which included sugar beet, cossettes, diffusion juice, second-phase diffusion juice, light juice and thick juice. The results showed that dominant genera included Pantoea, Pseudomonas, Leuconostoc and Burkholderia. Moreover, significant changes in bacterial communities were observed in samples. Regarding the relevant nitrogen metabolic potential, this study revealed communities with the ability for nitrate and nitrite metabolism. Furthermore, a shaking experiment involving diffusion juice and second-phase diffusion juice was performed, and results showed that the nitrate level declined 73% and 98% in 36 h, respectively. These results suggested that the bacterial communities contribute to nitrate and nitrite transformation. CONCLUSION: This study illustrated that the bacterial communities and their specific effects on the formation of nitrate and nitrite during beet sugar processing. The results presented the basic concept involving the nitrate- and nitrite-forming pathways directly related to the mechanism of bacterial community growth. This study could facilitate an understanding of the correlation between nitrite content and microorganisms to guide beet sugar manufacturers regarding the control of nitrite and nitrate content. © 2021 Society of Chemical Industry.

Characterizing the Volatile and Sensory Profiles, and Sugar Content of Beeswax, Beebread, Bee Pollen, and Honey.

Bee products are a well-known remedy against numerous diseases. However, from the consumers' perspective, it is essential to define factors that can affect their sensory acceptance. This investigation aimed to evaluate the volatile and sensory profiles, and sugar composition of beeswax, beebread, pollen, and honey. According to the HS-SPME/GC-MS results, 20 volatiles were identified in beeswax and honey, then 32 in beebread, and 33 in pollen. Alkanes were found to dominate in beeswax, beebread, and pollen, while aldehydes and monoterpenes in honey. In the case of sugars, a higher content of fructose was determined in beebread, bee pollen, and honey, whereas the highest content of glucose was assayed in beeswax. In the QDA, the highest aroma intensity characterized as honey-like and sweet was found in honey, while the acid aroma was typical of beebread. Other odor descriptors, including waxy, pungent, and plant-based aromas were noted only in beeswax, honey, and pollen, respectively.

Prospects of using hyperbranched stationary phase based on poly(styrene-divinylbenzene) in mixed-mode chromatography.

Evaluation of the chromatographic properties of covalently bonded hyperbranched stationary phase based on poly(styrene-divinylbenzene) (PS-DVB) and containing zwitterionic fragments in the structure of functional layer was conducted in suppressed ion chromatography (IC), reversed phase high performance liquid chromatography (RP HPLC), and hydrophilic interaction liquid chromatography (HILIC) modes. Besides the possibility of resolving 20 inorganic anions and organic acids using KOH eluent in suppressed IC, prepared resin provided the separation of alkylbenzenes in RP HPLC, water-soluble vitamins, amino acids, and sugars in HILIC mode. Trends in the retention of hydrophobic and polar analytes on the prepared stationary phase indicated the dominating effect of analyte nature on the retention mechanism and proved satisfactory hydrophilization of PS-DVB surface with hyperbranched functional layer for retaining polar compounds. The obtained results revealed good prospects of using hydrophobic PS-DVB substrate for preparing stationary phases for mixed-mode chromatography.

Characteristics of a recombinant Fusarium verticillioides cutinase and its effects on enzymatic hydrolysis of rice straw.

The current study heterologously expressed a cutinase from Fusarium verticillioides by Pichia pastoris and investigated its properties and effects on the hydrolysis of rice straw. The optimal pH and temperature for F. verticillioides cutinase were 8.0 and 50 °C, respectively. F. verticillioides cutinase had poor thermal stability and could be inhibited by some metal ions, inhibitors, and detergents (5 mM), including Ni2+, Zn2+, Cu2+, Ca2+, Mn2+, sodium dodecyl sulfate, EDTA, and Tween-20. F. verticillioides cutinase could tolerate 15% methanol and dimethyl sulfoxide but was significantly repressed by 15% ethanol and acetone with 48% and 63% residual activity, respectively. F. verticillioides cutinase could degrade the cuticle of rice straw with palmitic acid and stearic acid as the main products. However, the dissolving sugars released from the rice straw treated with F. verticillioides cutinase were significantly reduced by 29.2 µg/mL compared with the control (107.9 µg/mL). Similarly, the reducing sugars produced from the cellulase hydrolysis of rice straw pretreated with F. verticillioides cutinase were reduced by 63.5 µg/mL relative to the control (253.6 µg/mL). Scanning electron microscopy results showed that numerous tuberculate or warty protrusions were present nearly everywhere on the surface of rice straw treated with F. verticillioides cutinase, and some protrusions even covered and blocked the stomata of the rice straw surface. Current limited data indicate that F. verticillioides cutinase might not be an appropriate choice for improving the utilization of agricultural straws.

Is sugar extracted from plants less healthy than sugar consumed within plant tissues? The sugar anomaly.

There are dilemmas in the minds of consumers with respect to sugar consumption - they would like to consume sugars for sweetness, but in a healthy (and perhaps guilt free!) way. In a sense, consumers believe that if sugar does not appear as an ingredient on the product label, but is intrinsic in the food (and will appear as a nutrient), it is 'good'. As an ingredient, however, it is viewed as a 'bad chemical' associated with tooth decay and obesity. The reality is that unless processing induced modifications have occurred, the sugar molecule within a plant tissue is the same molecule structure as present in purified sugar. The same calorific value. However, there is an argument that humans eat too refined food and that if sugars were eaten in their natural context (e.g. within a fruit), their presence and concentration would be in harmony (where different nutrients complement and balance the sugar concentration) with the human body. This reflects the process of eating, satiety, presence of other nutrients (including water) and the associated impact of the indigestible components of plant foods on the transit/nutrient bioavailability control and thus benefits through the gut. The authors explore these issues in this article and seek to provide a scientific basis to different sides of the argument - sugar is good or bad depending on how (in which format and how much/how concentrated) it is consumed. More importantly perhaps, how should sugar consumption - an important nutrient - be managed to optimize the benefits but reduce the disadvantages? © 2020 Society of Chemical Industry.

Acute and chronic oral toxicity assessment of longan sugar extracts derived from whole fruit and from fruit pulp in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Longan (Dimocarpus longan Lour.) is one of the most popular subtropical fruits. Various parts of longan, including seeds, pericarp and pulp, have long been used in traditional medicine in China, Thailand and other Asian countries. The pulp has high sugar, vitamin and mineral content as well as bioactive components. The seeds and pericarp have also been reported to contain beneficial polyphenolic compounds. Longan sugar extract from pulp (LGSP) is prepared as a conventional sugar product. Longan sugar extract from whole longan fruit (LGSW) is also offered as a health food and as a medicinal product. AIM OF THE STUDY: The objective of this study was to identify and compare potential health hazards of both LGSW and LGSP by testing for acute and chronic oral toxicity in rats. MATERIALS AND METHODS: In acute toxicity testing, an oral dose (20 g/kg) of either LGSW or LGSP was administered to groups of rats. Mortality and clinical signs of toxicity were observed for 24 h, and then daily for a total of 14 days. In the chronic toxicity test, either LGSW (1, 2.5 and 5 g/kg/day) or LGSP (5 g/kg/day) was administered orally for a period of 180 days. After that treatment period, the rats in the satellite groups which received the highest doses of either LGSW or LGSP were observed for an additional 28 days. The rats then underwent clinical observation, body and organ weight measurement, hematological and biochemical analyses, and histopathological examination. RESULTS: In the acute toxicity study, the oral administration of LGSP or LGSW in either pellet or syrup formulations did not cause mortality or any pathological abnormalities. In the chronic toxicity study, neither LGSW nor LGSP resulted in death or in any changes in behavior of the rats. All hematological and serum biochemical values of both the LGSW- and LGSP-treated groups were within the normal ranges. No histopathological abnormalities of any internal organs were observed. CONCLUSION: The safety of longan sugar extract made from whole fruit (pulp, seeds and pericarb) is comparable to that of longan sugar extract made from pulp alone.

Improving the extraction and purification of leaf and phloem sugars for oxygen isotope analyses.

RATIONALE: The oxygen isotopic composition (here shown as the δ18 O value) of soluble sugars in leaves and phloem tissue holds valuable information about plant functions in response to climatic changes. However, δ18 O analysis of sugars is prone to error, and thoroughly tested methods are lacking. METHODS: We performed three experiments to test if sample preparation modifies the δ18 O values of sugars. In experiment 1, we tested the effects of oven-drying versus freeze-drying, whereas in experiment 2 we focused on the extraction and purification of leaf sugars. In experiment 3, we investigated the exudation and purification of twig phloem sugars as a function of exudation time and different ethylenediaminetetraacetic acid (EDTA) exudation media. RESULTS: Freeze-drying produced more consistent δ18 O values than oven-drying for sucrose but not for phloem sugars. The extraction and purification of leaf sugars can be performed without a significant modification of their δ18 O values; yet the purified leaf and phloem sugars possessed higher δ18 O values than the fraction of water-soluble compounds. Moreover, the exudation time significantly modulated the δ18 O values of phloem sugars, which is probably related to changes in the sugar composition. The addition of EDTA did not improve the determination of the δ18 O values of phloem sugars. CONCLUSIONS: We show that the sample preparation of plant sugars for the reliable determination of δ18 O values requires a strict protocol, which is described in this paper. For phloem sugar, we recommend a maximum exudation time of 1 h to reduce the degradation of sucrose and minimise oxygen isotope exchange reactions between the resulting hexoses and water.

Processing of coconut sap into sugar syrup using rotary evaporation, microwave, and open-heat evaporation techniques.

BACKGROUND: Coconut sugar has a caramel color with a taste like brown sugar. It is commonly used as natural sweetener. However, coconut sugar has been produced from coconut sap using a traditional method that involves heating the sap at high temperature (>100 °C) in an open pan for a long period (3-5 h). This conventional method results in an over-cooked sugar, which leads to quality deterioration in terms of both its physical and chemical properties. The current study aimed to investigate the processing of coconut sap into sugar syrup using alternative processing techniques such as rotary vacuum evaporation (RE) and microwave evaporation (ME), comparing them with open-heat evaporation (OHE) technique. RESULTS: Coconut sugar syrup produced by rotary evaporation at 60 °C and 250 mbar vacuum (RE-60) required the shortest production time (12.2 min) and the lowest processing temperature (54.8 °C) when compared with ME (13 min and 103.2 °C) and OHE (46.8 min and 101.6 °C). It also had a light brownish color with a higher L* value (35.17) than the ME (29.84) and OHE (23.84) methods. It was found to contain higher amounts of monosaccharides (fructose and glucose) and lower amounts of disaccharides (sucrose). Furthermore, the amount of energy required for RE-60 (0.35 kWh) was much less than for OHE (0.83 kWh). CONCLUSION: This study provided an alternative processing method for the sugar processing industry to produce coconut sugar using the rotary evaporation method at 60 °C under 250 mbar vacuum with better physicochemical qualities, shorter processing time, and minimum input energy. © 2020 Society of Chemical Industry.

Fourier-transform infrared imaging and multivariate analysis for direct identification of principal polysaccharides in brown seaweeds.

The current hydrocolloid industry requires new techniques for biomass characterization, which can quickly and ecologically characterize contained sugars. This work proposes the use of Fourier Transform Infrared microspectroscopy in combination with multivariate methods, to localize and identify the main carbohydrates and other components present in fresh brown seaweeds, avoiding time-consuming samples pre-treatments. Infrared images of Macrocystis pyrifera samples were analyzed by Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS) and Principal Component Analysis (PCA) as chemometrics techniques to identify the compounds. MCR-ALS was the best strategy, delivering pure spectra of chemical compound that PCA did not. The carbohydrates identified by this method were 1-3-ß-glucans divided into endofibers and laminarin; two types of fucoidans (rich in fucose or mannuronic acid), alginate and mannitol, besides other compounds such as proteins. This technique represents an opportunity for the hydrocolloid industry for a modern, rapid and environmentally-friendly characterization of macroalgal biomass to enhance its use.

Fractionation, structure and conformation characterization of polysaccharides from Anoectochilus roxburghii.

Two new isolated polysaccharides, ARPP-40 (40 % ethanol precipitate) and ARPP-70 (70 % ethanol precipitate), were extracted from Anoectochilus roxburghii. The physicochemical properties of two polysaccharides were analyzed and the results showed the relative weight average molecular weights and contents of neutral sugar for ARPP-40 and ARPP-70 were 423 kDa and 97.4 %, 10.8 kDa and 51.4 %, respectively. In terms of monosaccharide composition, ARPP-40 contained only glucose, while ARPP-70 was composed of seven monosaccharides, of which glucose and galactose were the main components. Furthermore, the structure and conformation characteristics of ARPP-40 were systematically investigated. The results revealed that ARPP-40 was supposed to be a glucan and existed as a flexible chain with a polydispersity index of 1.02 in 0.1 M NaNO3 solution. The systematic information on structural and conformational properties of ARPP-40 was meaningful for its further application in food and medicinal industry.

Profiling of volatile and non-phenolic metabolites-Amino acids, organic acids, and sugars of green tea extracts obtained by different extraction techniques.

Volatile compounds and non-phenolic metabolites (amino acids, organic acids, and sugars) of aqueous green tea extracts obtained by ultrasonic extraction (UE), agitation extraction (AE), hot water extraction (HWE), and conventional extraction (CE) were determined using SPME-GC-MS and HPLC, respectively. Significantly higher (P < 0.05) yields of volatiles and non-phenolic metabolites were obtained via UE and AE than via HWE and CE. UE, AE, HWE, and CE released 212, 201, 103, and 65 volatiles, respectively. Sum total of amino acid and organic acid in extracts was 54.57, 54.35, 27.11, and 12.67 (mg/100 g), and 5.96, 6.19, 3.81, and 1.68 (mg/100 g) for UE, AE, HWE, and CE, respectively. Volatiles except nitrogen-containing compounds had higher positive correlations with l-theanine, sucrose, malic acid, and catechins yields. Findings of the current study suggest that an efficient extraction technique may significantly increase volatile and non-phenolic metabolite yields in aqueous green tea extract.

High-energy ball milling treatment of soybean for Bacillus thuringiensis culture media.

Soybean meal has been intensively used as a substrate in culture media for several microorganisms. However, the fermentable sugar containing the soybean needs to be released from the solid matrix through different processes. Against this backdrop, the present study explores the use of high-energy ball milling as a one-step treatment method for expedited production of fermentable sugars of textured soybean. The best result is observed after only 5 min of milling, obtaining 34.1 times more fermentable sugars than untreated textured soybean, and 2.5 times more than commercially used soybean meal. Notably, the textured soybean ball-milled has been used as a substrate for Bacillus thuringiensis var. kurstaki HD-73 fermentation. The cell and spore production is also compared with a standard Rowe media. The maximum cell concentration obtained in the entire fermentation process using ball-milled textured soybean media is found to be higher than the concentration obtained using the standard Rowe media. In addition, it is observed that there is a direct correlation between maximum cell production and reducing sugar concentration generated by the high-energy ball milling treatment. No fermentation inhibitors or by-products are generated during the physical treatment.

Silver and gold nanoparticles based colorimetric assays for the determination of sugars and polyphenols in apples.

In this work, the exploitability of rapid and easy to use methods for the evaluation of the antioxidant capacity (AOC) and sugars content (SC), through metal nanoparticles (MNPs) formation, has been proved and applied to apples. In particular, an AgNPs-based sugar quantification assays and an AuNPs-based polyphenols antioxidant capacity assay have been used as tools for the evaluation of apple extracts composition. Both assays are based on the ability of the analytes (sugars and polyphenols) to reduce the source of metal (Au3+ and Ag+), stabilizing, at the same time, the resulting MNPs colloidal suspensions. The AuNPs and AgNPs formation depends on the analyte structure and concentration, resulting in red (AuNPs) and yellow (AgNPs) colored suspensions. Both assays require an initial mixing step, followed by MNPs formation under mild conditions (10 min, room temperature or 45 °C), and the colorimetric response is easily acquired at a fixed wavelength (AgNPs: 430 nm and AuNPs: 540 nm). The analytical performance of both assays has been proven, obtaining good reproducibility (RSD ≤ 6%,), sensitivity (LODs ≤8.7 µmol L-1) and recoveries (91% -113.7%). The produced MNPs (AgNPs and AuNPs) have been characterized using UV-Vis spectroscopy and TEM, and the cross-reactivity between assays, as well as the possible endogenous interferents, studied. The assays have been tested on 42 apple samples, and the data obtained compared with those obtained by conventional methods (i.e. FC, ABTS, and ion chromatography). The proposed AgNPs-sugars assay gives results comparable (R = 0.915) to those determined by ion chromatography in terms of total sugars, and the AuNPs-polyphenols assay results able to assess the polyphenols antioxidant capacity, being correlated with those obtained by the ABTS method (R = 0.922). This MNPs-based approach demonstrated to be an excellent tool for rapid and facile analysis of sugars and polyphenols in apple samples.

Two-Step Pretreatment of Corn Stover Silage Using Non-ionic Surfactant and Ferric Nitrate for Enhancing Sugar Recovery and Enzymatic Digestibility of Cellulose.

Corn stover silage (CSS) is regarded as a promising feedstock for bioethanol production. The two-step pretreatment using a sequential non-ionic surfactant and ferric nitrate pretreatment was investigated for improving the enzymatic hydrolysis of CSS. The first-step pretreatment using non-ionic surfactant (Tween-80, 2.0 wt.%) at 60 °C for 60 min achieved 30.48% the removal of lignin. Compared with the raw material, the cellulose content of first-step treated CSS increased by 15.86%. The second step using ferric nitrate resulted in 94.56% hemicellulose removal and achieved 72.53% cellulose purity at 130 °C for 30 min, while the yields of furfural and HMF were only 0.36 and 0.32 g/100 g dry material, respectively. The maximum enzymatic digestibility of the two-step treated CSS was 90.98% with a low cellulose dosage (15 FPU/g-glucan), which was approximately 32.07% higher than that of the first-step pretreatment only with Tween-80.

Separation of carbohydrates on electrophoresis microchips with controlled electrolysis.

This study reports the separation of fructose, galactose, glucose, lactose and sucrose on glass microchip electrophoresis (ME) devices using a microfluidic platform adapted with external reservoirs for controlling the electrolysis phenomenon. The connections between external reservoirs and microfluidic platform were performed by saline bridges created using silicone tubing filled with BGE. The separation conditions were optimized and the best results were achieved using a BGE containing 75 mmol/L NaOH and 15 mmol/L trisodium phosphate. Electrophoretic separations were monitored using a capacitively coupled contactless conductivity detection system. The controlled electrolysis has successfully allowed the application of a higher voltage on the separation channel promoting the baseline separation of five carbohydrates within 180 s with great run-to-run repeatability (RSD < 1%). The achieved efficiencies ranged from 45 000 ± 6000 to 70 000 ± 3000 plates/m demonstrating a performance better than ME devices without controlled electrolysis. The proposed system offered good linearity from 1 to 10 mmol/L and LODs between 150 and 740 µmol/L. The use of external tubes for controlling the electrolysis phenomenon on ME devices has solved common problems associated to run-to-run repeatability and analytical reliability required for routine and quantitative analysis.

Pressurized liquid extraction of cyclitols and sugars: optimization of extraction parameters and selective separation.

Cyclitols and sugars were obtained as a mixture from Medicago sativa L., in a comparative study by using maceration, and pressurized liquid extraction, as a modern and green extraction techniques. The influence of extraction parameters including: extraction temperature, time and number of cycles on the content of sugars and cyclitols was investigated based on response surface methodology. The highest total amount of sugars and cyclitols (62.27 ± 2.30 and 50.35 ± 0.77 mg/g of dry material, respectively) was obtained when extraction was performed at 88°C, for 22 min, in two cycles. The methodology used involved extraction, purification, selective separation (using yeast and anion exchange resin) and derivatization, followed by gas chromatography -mass spectrometry analysis. The use of yeast treatment realized an effective fractionation of cyclitols and sugars, which allowed the removal of most sugars. The involvement of anion exchange resin after yeast allowed the removal of sugar alcohols and lactose, together with other sugar traces remained and to obtain a solution containing six cyclitols. The recrystallization of dry residue after solvent evaporation, from ethanol, allowed us to obtain 14.65 mg of white pure crystals identified with NMR spectroscopy, liquid chromatography with mass spectrometry, gas chromatography with mass spectrometry, optical rotation and melting point as analysis D-pinitol.

Purification of the mother liquor sugar from industrial stevia production through one-step adsorption by non-polar macroporous resin.

Steviol glycosides (SGs)-enriched mother liquor sugar (MLS) used to be considered as a by-product of industrial stevia production. This work developed an efficient purification strategy to remove impurities from MLS. A non-polar macroporous adsorption resin (MAR) NDR-1 was selected as the adsorbent and ethanol solution was used as the mobile phase. The adsorption ratio of SGs and impurities was highly affected by ethanol concentration (v/v) due to the polarity difference and competition. In particular, in 25% (v/v) ethanol solution, SGs were mostly adsorbed on NDR-1 while impurities were completely dissolved in ethanol solution. After static adsorption in 25% ethanol solution and desorption in pure ethanol, the impurity content of MLS was decreased from 34.15% to 12.88%, with a high adsorption ratio of 92.24%. At the same condition, dynamic adsorption presented a better purification result with low impurity content of 4.78% and high SGs recovery rate of 89.88%.

Combined antioxidant-biofuel production from coffee silverskin.

Biorefinery concept asks for an integrated processing approach to exploit all biomass components. The self-sustainability target may be approached if molecules characterized by high added value and fermentable sugars are produced simultaneously. In the present study, sequential (i) mild hydrothermal pretreatment to produce antioxidants and (ii) NaOH pretreatment to produce a fermentable sugar solution were carried out on coffee silverskin. Twenty-minute treatment and biomass to liquid ratio 1:30 (g mL-1) were identified as optimal operating conditions to extract bioactive compounds characterized by antioxidant activity (22.2 mgGAE/gCSS; 13.9 mgTE/gCSS). Twenty-minutes and biomass to liquid ratio of 1:20 (g mL-1) were identified as optimal operating conditions to maximize sugar recovery and ABE production (solvent yield YABE/Sugars and ABE productivity of 0.21 g g-1 and 0.12 g L-1 h-1 were obtained, respectively). The study marks the highly economic potential of the process aimed to exploit the CSS as feesdstock for antioxidant and biofuel production.

Combined dilute hydrochloric acid and alkaline wet oxidation pretreatment to improve sugar recovery of corn stover.

Two-stage dilute hydrochloric acid (DA)/aqueous ammonia wet oxidation (AWO) pretreatment was used to recover the sugars of corn stover. The morphology characterizations of samples were detected by SEM, BET and SXT. The results showed that DA-AWO process demonstrated a positive effect on sugar recovery compared to AWO-DA. 82.8% of xylan was recovered in the first stage of DA-AWO process at 120 °C for 40 min with 1 wt% HCl. The second stage was performed under relative mild reaction conditions (130 °C, 12.6 wt% ammonium hydroxide, 3.0 MPa O2, 40 min), and 86.1% lignin could be removed. 71.5% of glucan was achieved with a low enzyme dosage (3 FPU·g-1) in the following enzymatic hydrolysis. DA-AWO pretreatment was effective due to its sufficient hydrolysis of hemicellulose in the first stage and remarkably removal of the lignin in the second stage, resulting in high sugar recovery with a low enzyme dosage.