Bioethanol production from sweet sorghum bagasse by Mucor hiemalis

The present work deals with production of ethanol from sweet sorghum bagasse by a zygomycetes fungus Mucor hiemalis. The bagasse was treated with phosphoric acid and sodium hydroxide, with or without ultrasonication, prior to enzymatic hydrolysis by commercial cellulase and β-glucosidase enzymes. The phosphoric acid pretreatment was performed at 50 °C for 30 min, while the alkali treatment performed with 12% NaOH at 0 °C for 3 h. The pretreatments resulted in improving the subsequent enzymatic hydrolysis to 79-92% of the theoretical yield. The best hydrolysis performance was obtained after pretreatment by NaOH assisted with ultrasonication. The fungus showed promising results in fermentation of the hydrolyzates. In the best case, the hydrolyzate of NaOH-ultrasound pretreated bagasse followed by 24 h fermentation resulted in about 81% of the corresponding theoretical ethanol yield. Furthermore, the highest volumetric ethanol productivity was observed in the hydrolyzates of NaOH pretreated bagasse, especially after ultrasonication in pretreatment stage.     

Optimization of formic/acetic acid delignification of Miscanthus × giganteus for enzymatic hydrolysis using response surface methodology

A Box-Behnken experimental design and response surface methodology were employed to optimize the pretreatment parameters of a formic/acetic acid delignification treatment of Miscanthus × giganteus for enzymatic hydrolysis. The effects of three independent variables, namely cooking time (1, 2 and 3 h), formic acid/acetic acid/water ratio (20/60/20, 30/50/20 and 40/40/20) and temperature (80, 90 and 107 °C) on pulp yield, residual Klason lignin content, concentration of degradation products (furfural and hydroxymethylfurfural) in the black liquor, and enzymatic digestibility of the pulps were investigated. The major parameter influencing was the temperature for pulp yield, delignification degree, furfural production and enzymatic digestibility. According to the response surface analysis the optimum conditions predicted for a maximum enzymatic digestibility of the glucan (75.3%) would be obtained using a cooking time of 3 h, at 107 °C and with a formic acid/acetic acid/water ratio of 40/40/20%. Glucan digestibility was highly dependent on the delignification degree.     

Optimization of ethanol production by Saccharomyces cerevisiae UFPEDA 1238 in simultaneous saccharification and fermentation of delignified sugarcane bagasse

Ethanol production by Saccharomyces cerevisiae UFPEDA1238 was performed in simultaneous saccharification and fermentation of delignified sugarcane bagasse. Temperature (32 °C, 37 °C), agitation (80; 100 rpm), enzymatic load (20 FPU/g cellulose and 10%, v/v β-glucosidase or 10 FPU/g cellulose and 5% β-glucosidase) and composition of culture medium were evaluated. Ethanol concentration, enzymatic convertibility of cellulose and volumetric productivity were higher than 25 g/L, 72% and 0.70 g/L h, respectively, after 30 h, when the culture medium 1 and 20 FPU/g cellulose/10%, v/v β-glucosidase or the culture medium 2 and 10 FPU/g cellulose/5% β-glucosidase were used in SSF at 37 °C and 80 rpm. In the SSF with culture medium 2 (supplemented with ammonium, phosphate, potassium and magnesium), 150 L ethanol/t bagasse was achieved, with minimum enzyme loading (10 FPU/g cellulose and 5%, v/v β-glucosidase) for 8%, w/v of solids, which is often an important requirement to provide cost-efficient second generation ethanol processes.     

Processing of Lespedeza stalks by pretreatment with low severity steam and post-treatment with alkaline peroxide

The steam pre-treatment with low severity preserves valuable biomass components, and further delignification with alkaline peroxide could improve hydrolysis. A combination of low severity steam pretreatment and alkaline peroxide post-treatment of Lespedeza stalks was investigated. The post-treatment of steam-pretreated Lespedeza stalks with alkaline peroxide significantly increased the cellulose content and changed the structure of the cellulose-rich fractions. A glucose yield of 503.5 mg g⁻¹ raw material from enzyme hydrolysis was obtained when the steam-pretreated material (184 °C for 4 min) was post-treated with 2% hydrogen peroxide at 60 °C for 24 h with a substrate concentration of 3.3%. Its hydrolysis yield is 88.8%, which is higher than that of samples processed by steam pretreatment alone (63.7%). The samples obtained by post-treatment with alkaline peroxide were found to have a smoother surface and looser structure in scanning electron microscopy images. The isolated lignin preparations had a yield range from 10.9 to 14.7 (% dry matter). The lignin was characterized by thermogravimetric analysis/differential thermal analysis, Fourier transform infrared spectroscopy, and gel permeation chromatography. Alkaline peroxide treatment increased the thermal stability of lignin, and decreased the amounts of all functional groups. Depolymerization and repolymerization occurred during the alkaline peroxide treatment.     

Utilization of pretreated bagasse for the sustainable bioproduction of cellulase by Aspergillus nidulans MTCC344 using response surface methodology

Response surface methodology (RSM) was used to optimize the conditions for the production of endo β-1,4 glucanase, a component of cellulase by Aspergillus nidulans MTCC344 under solid state fermentation, using bagasse as the chief substrate. A four-factor-five-level central composite design was employed for experimental design and analysis of the results. Maximum cellulase activity (CMCase was 28.96 U g⁻¹) can be attained at the optimum conditions, 16.8 mm bagasse bed height, 60% moisture content, pH 4.25 and temperature 40 °C in the solid state fermenter. These data were rather close to the experimental results obtained (CMCase was 28.84 U g⁻¹). A. nidulans MTCC344 was able to hydrolyze pretreated bagasse completely after 8 days of incubation with significant endo β-1,4 glucanase activities. The results of Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and scanning electron microscopy (SEM) of bagasse showed structural changes through pretreatment, in favor of enzymatic hydrolysis. Bagasse with alkali pretreatment using sodium hydroxide is a source of lignocelluloses able to improve the yield of endo β-1,4 glucanase by the strain of A. nidulans. The endo β-1,4 glucanase produced during the bioconversion of cellulose to glucose by A. nidulans MTCC344 is strongly dependent on the pretreatment given before hydrolysis.     

Ethanol production from alkali- and ozone-treated cotton stalks using thermotolerant Pichia kudriavzevii HOP-1

In the present study, milled cotton stalks were subjected to alkali pretreatment with NaOH at 1-4% (w/v) concentrations at 121 °C for time ranging from 30 to 90 min. Ozone pretreatment was performed by passing 45 mg/L of ozone gas over 2 mm cotton stalks for 150 min at a flow rate of 0.37 L/min. The residual biomass from 4% alkali pretreatment for 60 min showed 46.6% lignin degradation accompanied by 83.2% increase in glucan content, compared with the untreated biomass. Hydrolysis of 4% alkali-treated and ozone-treated cotton stalks was conducted using enzyme combination of 20 filter paper cellulase units/gram dried substrate (FPU/g-ds), 45 IU/g-ds β-glucosidase and 15 IU/g-ds pectinase. Enzymatic hydrolysis of alkali-treated and ozone-treated biomass after 48 h resulted in 42.29 g/L glucose, 6.82 g/L xylose and 24.13 g/L glucose, 8.3 g/L xylose, respectively. About 99% of glucose was consumed in 24 h by Pichia kudriavzevii HOP-1 cells resulting in 19.82 g/L of ethanol from alkali-treated cotton stalks and 10.96 g/L of ethanol from ozone-treated cotton stalks. Simultaneous saccharification and fermentation of the alkali-treated cotton stalks after 12-h pre-hydrolysis resulted in ethanol concentration, ethanol yield on dry biomass basis and ethanol productivity of 19.48 g/L, 0.21 g/g and 0.41 g/L/h, respectively which holds promise for further scale-up studies. To the best of our knowledge, this is the first study employing SSF for ethanol production from cotton stalks.     

Preparation of shikonin by hydrolyzing ester derivatives using basic anion ion exchange resin as solid catalyst

An efficient catalytic hydrolysis approach was developed to prepare shikonin from its ester derivatives with anion exchange resins as solid catalyst. The performance of seven anion exchange resins including D290, D296, D261, D280, 201×7, D301-G and D301-R has been evaluated. The research results indicate that D290 resin is the most appropriate for preparation of shikonin. The reaction conditions such as reaction temperature, reaction time, catalyst amount and catalyst reuse have been studied. The maximum yield of shikonin reached 93.74 ± 2.99% under optimal conditions of hydrolysis temperature 20 °C, hydrolysis and desorption time both 30 min and resin amount 10% (w/v). HPLC chromatograms of samples, obtained after the new catalytic process, demonstrated that conversion of shikonin ester derivatives into shikonin performed well.     

Extraction of flax shive using sodium ethoxide catalyst in anhydrous ethanol

This work investigated the yield and nature of solvent-soluble organic compounds extracted from flax shive using a room temperature reaction (20 °C) with sodium ethoxide catalyst at four different concentrations (0.2, 0.5, 0.7, and 1.0 M) in anhydrous ethanol. Results were compared with the use of aqueous sodium hydroxide (1.0 M) at two different reaction temperatures (20 °C and 100 °C). Quantitative yield from flax shive varied linearly with sodium ethoxide concentration and averaged 54.5 mg/g on a dry-mass basis (db) at 1.0 M. In contrast, the quantitative yield using 1.0 M sodium hydroxide was much lower, averaging 2.2 mg/g (db). Yield did not differ significantly due to changes of particle size in either case, or due to changes of temperature over the range considered in the case of sodium hydroxide.Analyses using proton nuclear magnetic resonance (¹H NMR) confirmed all extracts to contain aromatic compounds, thus likely lignin derived, but found differences in chemical characteristics between the two extraction methods. One key difference was the presence of compounds with methyl ether groups in sodium hydroxide extracts that were absent in the case of sodium ethoxide extracts. Given that flax contains a mixed guaiacyl-syringyl lignin, methyl ether groups would be expected to be present. Control reactions on three model compounds were carried out to confirm that transesterification occurred with sodium ethoxide. These control reactions also demonstrated that methyl ether groups would be expected to remain intact under the extraction conditions reported here. In light of the higher yield of solvent soluble compounds recovered by extraction with basic ethanol, flax shive may represent a source of value-added phenolic constituents. This processing method may also represent a useful pre-treatment prior to the production of biofuels by cellulose degrading organisms.     

Optimization of ethanol production from microwave alkali pretreated rice straw using statistical experimental designs by Saccharomyces cerevisiae

Ethanol production from agro-waste provides an alternative energy-production system. Statistical experimental designs were used for optimization of critical nutrients and process variables for ethanol production. The critical nutrients and process variables were initially selected according to a Placket-Burman (PB) design. Selected factors (inoculum level 1-5%, pH 4.5-7, temperature 25-35 °C and urea concentration 0.25-0.75 g/L) were optimized by response surface methodology (RSM) based on a three-level four-factor Box-Behnken design (BBD). Under optimum conditions of inoculum level 3%, pH 5.75, temperature 30 °C and urea concentration 0.50 g/L maximum ethanol production obtained 13.2 g/L from microwave alkali pretreated rice straw with ethanol productivity 0.33 g/L/h. Under optimum conditions ethanol production studied at fermenter level and obtained ethanol concentration 19.2 g/L, ethanol productivity 0.53 g/L/h and ethanol yield to consumed sugar 0.50 g/g. These results indicated that ethanol production can be enhanced by optimization of nutritional and process variables.     

Characterization of depolymerized residues from extremely low acid hydrolysis (ELA) of sugarcane bagasse cellulose: Effects of degree of polymerization, crystallinity and crystallite size on thermal decomposition

Sugarcane bagasse cellulose was subjected to the extremely low acid (ELA) hydrolysis in 0.07% H₂SO₄ at 190, 210 and 225 °C for various times. The cellulose residues from this process were characterized by TGA, XRD, GPC, FTIR and SEM. A kinetic study of thermal decomposition of the residues was also carried out, using the ASTM and Kissinger methods. The thermal studies revealed that residues of cellulose hydrolyzed at 190, 210 and 225 °C for 80, 40 and 8 min have initial decomposition temperature and activation energy for the main decomposition step similar to those of Avicel PH-101. XRD studies confirmed this finding by showing that these cellulose residues are similar to Avicel in crystallinity index and crystallite size in relation to the 110 and 200 planes. FTIR spectra revealed no significant changes in the cellulose chemical structure and analysis of SEM micrographs demonstrated that the particle size of the cellulose residues hydrolyzed at 190 and 210 °C were similar to that of Avicel.     

Sugarcane bagasse whiskers: Extraction and characterizations

This work evaluates the use of sugarcane bagasse (SCB) as a source of cellulose to obtain whiskers. These fibers were extracted after SCB underwent alkaline peroxide pre-treatment followed by acid hydrolysis at 45 °C. The influence of extraction time (30 and 75 min) on the properties of the nanofibers was investigated. Sugarcane bagasse whiskers (SCBW) were analyzed by transmission electron microscopy (TEM), X-ray diffraction (XRD) and thermogravimetric analysis (TGA) in air atmosphere. The results showed that SCB could be used as source to obtain cellulose whiskers and they had needle-like structures with an average length (L) of 255 ± 55 nm and diameter (D) of 4 ± 2 nm, giving an aspect ratio (L/D) around 64. More drastic hydrolysis conditions (75 min) resulted in less thermally stable whiskers and caused some damage on the crystal structure of the cellulose as observed by XRD analysis.     

Comprehensive utilization of the hydrolyzed productions from rice hull

The study based on pretreatment, hydrolyzation and separation processes with the raw material rice hull, provides a comprehensive utilization of the hydrolyzed productions, such as glucose (C₆H₁₂O₆) from cellulose, silica (SiO₂), and byproduct crystalline sodium sulfate (Na₂SO₄·10H₂O). The optimum hydrolysis conditions are as follows: the concentration of H₂SO₄ is 72% (wt.%), the temperature is 50 °C, the ratio of H₂SO₄ solution volume (mL) to the rice hull mass (g) is 10:1 and the time is 5 min, the glucose yield rate reaches 45.6% (wt.%). The concentration of glucose solution could be 0.1 g/mL after neutralization measured by ultraviolet spectrophotometer (UV-VIS). Silica powder was below 50 nm characterized by X-ray diffraction (XRD) and scanning electron microscopy (SEM). The main byproduct crystalline sodium sulfate was featured by XRD and photographs.     

Kinetic modeling of enzymatic saccharification using wheat straw pretreated under autohydrolysis and organosolv process

The enzymatic saccharification kinetics of untreated wheat straw, pretreated solids obtained by a sequence of autohydrolysis (solubilization of hemicellulose) and organosolv (solubilization of lignin) were studied together with two pure cellulose model substrates, filter paper and Avicel. Two kinetic models for glucose production were compared and its kinetic constants calculated. According to the obtained results, enzymatic saccharification of the autohydrolysis pretreated solids (APS) proved to be more effective than when the organosolv pretreated solids (OPS) were used. The maximum extent of the enzymatic conversion of cellulose to glucose was 90.88% and 64.04%, for APS and OPS respectively, at 96 h. This result was probably due to an increase in accessible area for APS and a possible inhibition by phenolic acids deposited on the surface of OPS, acting as a barrier for enzymatic saccharification. Initial saccharification rate for APS and OPS was 0.47 g/(L h) and 0.34 g/(L h), respectively. Models based on first and second order cellulase deactivation kinetics satisfactory predicted the behavior of glucose production, however the second order model had a higher accuracy than the first order one. Visualization of structural modification induced by enzymatic saccharification at 12 h for the pretreated solids was done using scanning electron microscopy.     

Development of transparent bacterial cellulose nanocomposite film as substrate for flexible organic light emitting diode (OLED) display

Nanocomposite film composed of bacterial cellulose (10-50 wt.%) and poly-urethane (PU) based resin was fabricated and utilized as a substrate for flexible organic light emitting diode (OLED) display. The performance of the nanocomposite satisfied the criteria for the substrate of OLED with an additional feature of flexibility. The visible light transmittance of the nanocomposite film was as high as 80%. Its thermal stability was stable up to 150 °C while its dimensional stability in terms of coefficient of thermal expansion (CTE) was less than 20 ppm/K. After OLED was fabricated on the substrate through thermal evaporation technique, the OLED performed highest current efficiency of 0.085 cd/A and power efficiency of 0.021 lm/W at 200 cd/m² while retained its flexible feature, suggesting that bacterial cellulose nanocomposite is a promising material for the development of substrate for flexible OLED display.     

Fractionation of flax shives with pressurized aqueous ethanol

Fractionation of flax shives into major biopolymer constituents, such as cellulose, hemicelluloses, and lignin, was carried out with pressurized aqueous ethanol in a pressurized low-polarity water extractor. The effect of processing parameters such as temperature, ethanol concentration, flow rate, sample size and solvent/feed ratio on the simultaneous extraction of hemicelluloses and lignin was determined. More than 80% of total hemicelluloses and ∼78% of total lignin were removed simultaneously in a single step under the following conditions: 180 °C, 30% (v/v) ethanol concentration, 3 mL/min flow rate, and 45 mL/g solvent/feed ratio. Under these extraction conditions, cellulose degradation was negligible. Further, the separation of lignin from hemicelluloses was carried out using two simple alternative methodologies based on precipitation. Since no acidic or alkali catalysts were used, the degradation of biopolymers was negligible and the oligomer/monomer ratio of sugars was 825:1. Characterization of fractionated biopolymers was carried out with scanning electron microscopy and a Fourier-transform infrared spectrometer (FT-IR). FT-IR spectra of isolated lignin and hemicelluloses showed that both polymers were comparable to commercially available products.     

Optimization of switchgrass and extruder parameters for enzymatic hydrolysis using response surface methodology

Switchgrass is an important biomass that can be hydrolyzed to yield fermentable sugars through pretreatment, which is the primary and expensive step in conversion of biomass to bio-ethanol. Most of the pretreatment operates in batch mode, which is energy intensive, requires high capital, results in decomposition of hemicellulose, and formation of inhibitors. Considering these shortcomings, a novel biomass pretreatment method using a high shear bioreactor could be a viable continuous one. The current study was undertaken to determine the effect of biomass parameters such as moisture content (10, 20, 30, 40, and 50% wb) and particle size (2, 4, 6, 8, and 10 mm) over a range of barrel temperature and screw speed (45-225 °C and 20-200 rpm). Statistical analyses revealed that among the independent variables considered temperature, screw speed, and moisture content had significant effect on sugar recoveries. Proposed quadratic model to predict glucose, xylose, and combined sugar recoveries from switchgrass had a high F and R² values indicating that the model has the ability to represent the relationship among the independent variables studied. The optimum pretreatment condition of barrel temperature 176 °C, screw speed 155 rpm, moisture content 20% wb, and particle size 8 mm resulted in maximum glucose, xylose, and combined sugar recoveries of 41.4, 62.2 and 47.4%, respectively. The optimum pretreated switchgrass had 50% higher surface area than that of the control.     

Optimization of phosphoric acid catalyzed fractionation and enzymatic digestibility of flax shives

Flax shives are the woody residue left over from processing flax straw into fiber, and are an abundant renewable lignocellulosic material with a potential for the conversion into bioethanol and other value added products. In this study, prior to enzymatic hydrolysis for the liberation of fermentable sugars, such as glucose and xylose, flax shives were treated with concentrated phosphoric acid. In order to optimize the phosphoric acid pretreatment and enzymatic hydrolysis steps, the effects of three process variables on the fractionation of flax shives, and enzymatic digestibility of pretreated flax shives were evaluated. The optimization process employed a central composite design (CCD), where the variables selected were concentration of phosphoric acid (40.8–86.2%), pretreatment time (9.5–110.5 min), and cellulase loading (13.1–71.9 FPU/g cellulose). Using three-variable and five-level CCD, all tested independent variables were identified to have significant effects (P < 0.05) on the digestibility of pretreated flax shives. It was found that the level of phosphoric acid (P < 0.0001) affects the digestibility most significantly when compared with other variables. When the optimization was conducted under a constrain of minimum cellulase loading, the maximum digestibility of 94.8% was predicted when the phosphoric acid concentration, pretreatment time, and cellulase loading were 86.2%, 110.5 min, and 13.1 FPU/g cellulose at 50 °C and 120 h, respectively. Under these conditions, digestibility of pretreated flax shives in the validation study reached a maximum of 93% at 120 h of incubation, showing good agreement with the values from the validation experiment of 93.4%, indicating high accuracy of the CCD procedure. When triticale straw, pine wood, and poplar wood were pretreated and hydrolyzed under optimum conditions obtained from the flax shives experiment, the digestibility reached 98.2, 74.8, and 95.7%, respectively, suggesting that the modest pretreatment process using phosphoric acid is an effective method for perennial plants as well as hard wood.     

Thermogravimetric analysis and the optimisation of bio-oil yield from fixed-bed pyrolysis of rice husk using response surface methodology (RSM)

The effects of pyrolysis temperature, heating rate, particle size, holding time, and gas flow rate were investigated to optimize bio-oil yield from rice husk pyrolysis. Thermogravimetric analysis showed thermal degradation of hemicellulose, cellulose and lignin, indicating faster decomposition of cellulose compared to lignin. The optimisation process was analysed by employing central composite design (CCD) in response surface methodology (RSM) using Design Expert Version 7.5.1 (StatEase, USA). A two-level fractional factorial was initially carried out and followed by RSM. The statistical analysis showed that pyrolysis temperature, heating rate, particle size and holding time significantly affected the bio-oil yield. By utilising response surface method, these four factors were investigated, analysed and optimal conditions were obtained at pyrolysis temperature of 473.37 °C, heating rate of 100 °C/min, particle size of 0.6 mm and holding time of 1 min. Confirmation runs gave 48.30% and 47.80% of bio-oil yield compared to 48.10% of predicted value. Furthermore, the pyrolytic bio-oils obtained from fixed-bed pyrolysis were examined using gas chromatographic/mass spectroscopy (GC/MS), Fourier transform infrared (FTIR) methods, elemental analyzer, pH probe and bomb calorimeter.     

Supercritical carbon dioxide (SC-CO2) extraction of Nigella sativa L. oil using full factorial design

In this study, Nigella sativa L. oil was extracted using supercritical carbon dioxide with full factorial design to determine the best extraction condition (pressure, temperature and dynamic extraction time) for obtaining an extract with high yield, antioxidant activity and thymoquinone (TQ) quantity. The maximum thymoquinone content in the highest overall yield was achieved through SC-CO₂ extraction condition of 150 bar, 40 °C, 120 min with the value of 4.09 mg/ml. The highest SC-CO₂ extraction yield was 23.20% which obtained through extraction condition of 350 bar, 60 °C and 120 min. The extraction conducted at 350 bar, 50 °C, 60 min showed the lowest IC₅₀ value (highest antioxidant activity) of 2.59 mg/ml using DPPH radical scavenging activity method. Fatty acid composition of the extracted oil with highest radical scavenging activity was obtained by gas chromatographic analysis.     

Conversion of starch from dry common beans (Phaseolus vulgaris L.) to ethanol

Dry common beans (Phaseolus vulgaris L.) were evaluated for potential conversion of starch to ethanol. Eight varieties of beans with average starch content of 46% (db) were assayed in a laboratory-scaled process based upon the commercial corn dry grind fermentation process. Ethanol yield was 0.43-0.51 g ethanol/g glucose (0.19-0.23 g ethanol/g beans). The average ethanol yield for the eight bean types was 92% of maximum theoretical yield, demonstrating that starch from beans could be efficiently converted to ethanol. Ethanol concentration obtained from 20% (w/w) solids loading was 3.5-4.4% (w/v). The residual fermentation solids contained, on a dry basis, 37.1-43.6% crude protein, 10.8-15.1% acid detergent fiber and 19.1-31.3% neutral detergent fiber.