Combined ethanol and methane production from switchgrass (Panicum virgatum L.) impregnated with lime prior to steam explosion

Pretreatments are crucial to achieve efficient conversion of lignocellulosic biomass to soluble sugars. In this light, switchgrass was subjected to 13 pretreatments including steam explosion alone (195 °C for 5, 10 and 15 min) and after impregnation with the following catalysts: Ca(OH)₂ at low (0.4%) and high (0.7%) concentration; Ca(OH)₂ at high concentration and higher temperature (205 °C for 5, 10 and 15 min); H₂SO₄ (0.2% at 195 °C for 10 min) as reference acid catalyst before steam explosion. Enzymatic hydrolysis was carried out to assess pretreatment efficiency in both solid and liquid fraction. Thereafter, in selected pretreatments the solid fraction was subjected to simultaneous saccharification and fermentation (SSF), while the liquid fraction underwent anaerobic digestion (AD). Lignin removal was lowest (12%) and highest (35%) with steam alone and 0.7% lime, respectively. In general, higher cellulose degradation and lower hemicellulose hydrolysis were observed in this study compared to others, depending on lower biomass hydration during steam explosion. Mild lime addition (0.4% at 195 °C) enhanced ethanol in SSF (+28% than steam alone), while H₂SO₄ boosted methane in AD (+110%). However, methane represented a lesser component in combined energy yield (ethanol, methane and energy content of residual solid). Mild lime addition was also shown less aggressive and secured more residual solid after SSF, resulting in higher energy yield per unit raw biomass. Decreased water consumption, avoidance of toxic compounds in downstream effluents, and post process recovery of Ca(OH)₂ as CaCO₃ represent further advantages of pretreatments involving mild lime addition before steam explosion.     

Physicochemical characterization and enzymatic digestibility of Chinese pennisetum pretreated with 1-ethyl-3-methylimidazolium acetate at moderate temperatures

1-ethyl-3-methylimidazolium acetate ([EMIM] AC) pretreatment at moderate temperatures (60 °C and 75 °C) was evaluated for improving hydrolysability of Chinese pennisetum, a leading candidate as an energy crop, for bioethanol production. The pretreatment caused slight carbohydrate and lignin loss but significantly changed the material physicochemical characters, such as crystallinity and surface structure. Both changes exhibited positive effects on improving the enzymatic digestibility of the Chinese pennisetum. It was observed that approximately 90% of the cellulose and 50% of the xylan in the Chinese pennisetum after pretreatment at 75 °C were converted to fermentable monosaccharides by the combined cellulases and endo-xylanase. The results suggested that Chinese pennisetum could be effectively pretreated with ([EMIM] AC) pretreatment at moderate temperatures, and the high hydrolysis yield of fermentable sugars from pretreated Chinese pennisetum could be achieved by the synergistic action of accessory xylanase in enzymatic hydrolysis by cellulases.     

Improvement of enzymatic saccharification of Populus and switchgrass by combined pretreatment with steam and wet disk milling

To reduce the recalcitrance of lignocellulosic biomass for subsequent biological processing, we pretreated energy crop feedstocks with mild steam treatment (ST; 130 and 150 °C for 60 min) and wet disk milling (WDM). We tested two phylogenetically different, but typical energy crop feedstocks: Populus trichocarpa and switchgrass (Panicum virgatum). WDM after ST facilitated the fibrillation of both types of biomass, resulting in an increase of specific surface area, improved enzymatic saccharification yield, and decrease in cellulose crystallinity. After steam treatment at 150 °C followed by 17 cycles of WDM, enzymatic hydrolysis resulted in almost complete glucan to glucose conversion in both feedstocks.     

Sweet sorghum as a whole-crop feedstock for ethanol production

The potential of sweet sorghum as an alternative crop for ethanol production was investigated in this study. Initially, the enzymatic hydrolysis of sorghum grains was optimized, and the hydrolysate produced under optimal conditions was used for ethanol production with an industrial strain of Saccharomyces cerevisiae, resulting in an ethanol concentration of 87 g L⁻¹. From the sugary fraction (sweet sorghum juice), 72 g L⁻¹ ethanol was produced. The sweet sorghum bagasse was submitted to acid pretreatment for hemicellulose removal and hydrolysis, and a flocculant strain of Scheffersomyces stipitis was used to evaluate the fermentability of the hemicellulosic hydrolysate. This process yielded an ethanol concentration of 30 g L⁻¹ at 23 h of fermentation. After acid pretreatment, the remaining solid underwent an alkaline extraction for lignin removal. This partially delignified material, known as partially delignified lignin (PDC), was enriched with nutrients in a solid/liquid ratio of 1 g/3.33 mL and subjected to simultaneous saccharification and fermentation (SSF) process, resulting in an ethanol concentration of 85 g L⁻¹ at 21 h of fermentation. Thus, from the conversion of starchy, sugary and lignocellulosic fractions approximately 160 L ethanol.ton⁻¹ sweet sorghum was obtained. This amount corresponds to 13,600 L ethanol.ha⁻¹.     

Evaluation of dilute acid and alkaline pretreatments,enzymatic hydrolysis and fermentation of napiergrass for fuel ethanol production

Napiergrass (Pennisetum purpureum Schum.) is a promising low cost raw material which does not compete with food prices, has attractive yields and an environmentally friendly farming. Dilute sulfuric acid pretreatment of napiergrass was effective to obtain high yields of sugars and low level of degradation by-products from hemicellulose. Detoxification with Ca(OH)₂ removed inhibitors but showed sugars loss. An ethanol concentration of 21 g/L after 176 h was found from the hydrolyzate using Pichia stipitis NBRC 10063 (fermentation efficiency 66%). An additional alkaline pretreatment applied to the solid fraction remaining from the diluted acid pretreatment improved the lignin removal. The highest cellulose hydrolysis values were found with the addition of β-glucosidase and PEG 6000. The simultaneous hydrolysis and fermentation of the cellulosic fraction with Saccharomyces cerevisiae, 10% (w/v) solid concentration, β-glucosidase and PEG 6000, showed the highest ethanol concentration (24 g/L), and cellulose hydrolysis values (81%). 162 L ethanol/t of dry napiergrass were produced (overall efficiency of 52%): 128 L/t from the cellulosic fraction and 34 L/t from the hemicellulosic fraction.     

Structural and chemical analysis of process residue from biochemical conversion of wheat straw (Triticum aestivum L.) to ethanol

Biochemical conversion of lignocellulose to fermentable carbohydrates for ethanol production is now being implemented in large-scale industrial production. Applying hydrothermal pretreatment and enzymatic hydrolysis for the conversion process, a residue containing substantial amounts of lignin will be generated. So far little is known about the composition of this lignin residue which at present is mainly incinerated for heat and power generation and not yet converted so much into more valuable products.In this study, the structural and chemical composition of the solid and liquid fractions of lignin residue from wheat straw were analysed and processing factors discussed. Roughly 70 and 15% of the solid mass fraction consisted of lignin and ash, respectively. Residual carbohydrates mostly originated from hemicellulose in the liquid fraction and from cellulose in the solid fraction. The solid fraction also contained significant amounts of protein, which is a valuable by-product when used as animal feed or when enzymes and yeast cells are separated for process recycling. Silica was the dominant constituent in the mineral fraction and except for few fragments of lignified middle lamellae most particles in the solid fraction appeared as silica coated by lignin, hampering separation of the two components before incineration or refinement of the residue.     

Energy recovery from grass of urban roadside verges by anaerobic digestion and combustion after pre-processing

Grass from urban roadside verges is a potential, though widely unused, resource for bioenergy recovery. Two possible bioenergy recovery techniques were tested, i.e. i) direct anaerobic digestion of the whole parent material and ii) the “integrated generation of solid fuel and biogas from biomass” (IFBB) procedure, which divides biomass into a press fluid and a press cake by mashing and mechanical dewatering. Biomass yield, chemical composition and canopy height of biomass, contribution of functional groups, fermentation characteristics of silage and press fluids, as well as characteristics of the produced solid fuel was investigated, applying a 4-cut management for anaerobic digestion, a 2-cut management for IFBB and an 8 times mulching as a reference. Mean annual biomass yield (2013 and 2014) was 3.24, 3.33 and 5.68 t dry matter ha⁻¹ for the mulching, 4-cut management and 2-cut management, respectively. Yields were higher in 2014 due to more favourable weather conditions. Fibre concentration was higher in material of the 2-cut management than in the 4-cut management, however, methane yield of the corresponding silages was the same. Highest methane yield was gained from press fluids with 292 l_N kg⁻¹ volatile solids. The press cake had a lower heating value of 16 MJ kg⁻¹ dry matter and a K₂O/CaO index of 0.51–0.88. Gross energy output was 26.4 GJ ha⁻¹ for anaerobic digestion and 84.4 GJ ha⁻¹ for IFBB. Thus, an altered roadside verge management with reduced cutting frequency might allow a significant energy recovery and improved ecosystem services, i.e. increased biodiversity.     

Corn stover ethanol yield as affected by grain yield,Bt trait,and environment

Literature values for glucose release from corn stover are highly variable which would likely result in tremendous variability in bio-refinery ethanol yield from corn stover feedstock. A relatively recent change in United States corn genetics is the inclusion of the Bacillus thuringiensis (Bt) trait, which now accounts for three-fourths of all US planted corn acreage. The objective of this study was to evaluate the effect of corn grain yield, inclusion of the Bt trait, and location environment on corn stover quality for subsequent ethanol conversion. Two hybrid pairs (each having a Bt and non-Bt near-isoline) were analyzed giving a total of 4 hybrids. In 2010 and 2011, field plots were located in Michigan at four latitudinal differing locations in four replicated plots at each location. Stover composition and enzymatic digestibility was analyzed and estimated ethanol yield (g g⁻¹) was calculated based on hydrolyzable glucan and xylan levels. Analysis showed that there were no significant differences in total glucose or xylose levels nor in enzymatically hydrolyzable glucan and xylan concentrations between Bt corn stover and the non-Bt stover isolines. Regression analyses between corn grain yield (Mg ha⁻¹) and corn stover ethanol yield (g g⁻¹) showed an inverse relationship indicative of a photosynthate source-sink relationship. Nevertheless, the quantity of stover produced was found to be more critical than the quality of stover produced in maximizing potential stover ethanol yield on a land area basis.     

Enhanced substrate degradation and methane yield with maleic acid pre-treatments in biomass crops and residues

Organic acids are envisaged as alternative catalysts to strong mineral acids, in pre-treatment of ligno-cellulosic biomass for anaerobic digestion (AD). To evaluate this hypothesis, an untreated control and four pre-treatments (25 °C for 24 h) involving two levels of maleic acid (34.8 and 69.6 kg m⁻³), alone and combined with sulphuric acid (4 kg m⁻³), were studied in three agricultural substrates: Arundo (aka giant reed), Barley straw and B133 fibre sorghum. Methane production was assessed in a batch AD assay (35 °C for 51 days) with 4 g L⁻¹ of volatile solid (VS) load. Fibre composition and structure were investigated through chemical analysis and Fourier transform infrared (FTIR) spectrometry. Arundo and B133 that were the most and least recalcitrant substrate, respectively, staged the highest and lowest increase in methane with high maleic acid: +62% over 218 cm³ g⁻¹ of VS in untreated Arundo; +36% over 284 cm³ g⁻¹ of VS in untreated B133. Barley straw showed an intermediate behaviour (+41% over 269 cm³ g⁻¹ of VS). H₂SO₄ addition to maleic acid did not improve CH₄ output. The large increase in methane yield determined by pre-treatments was reflected in the concurrent decrease of fibre (between 14 and 39% depending on fibrous component). Based on FTIR spectra, bands assigned to hemicellulose and cellulose displayed lower absorbance after pre-treatment, supporting the hypothesis of solubilisation of structural carbohydrates and change in fibre structure. Hence, maleic acid was shown a suitable catalyst to improve biodegradability of ligno-cellulosic biomass, especially in recalcitrant substrates as Arundo.     

Steam explosion pretreatment and enzymatic saccharification of duckweed (Lemna minor) biomass

Our previous research has shown that duckweed is potentially an ideal feedstock for the production of biofuels because it can be effectively saccharified enzymatically. Here we report the results of experiments in which duckweed was pre-treated by steam explosion prior to enzyme digestion. A range of temperatures, from 130 to 230 °C with a fixed retention time of 10 min, were employed. The best pretreatment conditions were 210 °C for 10 min; these conditions produced the highest amount of water-soluble material (70%), the greatest levels of starch solubilisation (21%) and hemicellulose and pectic polysaccharides degradation (60%). The use of these steam explosion conditions enabled large reductions in the concentrations of enzymes required for effective saccharification. The amount of Celluclast required was reduced from 100 U (4.35 FPU) g⁻¹ substrate to 20 U g⁻¹ substrate, and additional beta-glucosidase was reduced from 100 to 2 U g⁻¹ substrate.     

Improving methane production from wheat straw by digestate liquor recirculation in continuous stirred tank processes

Wheat straw is an abundant, cheap substrate that can be used for methane production. However, the nutrient content in straw is inadequate for methane fermentation. In this study, recycling digestate liquor was implemented in single-stage continuous stirred tank processes for enrichment of the nutrient content of straw with the aim of improving the methane production. The VS-based organic loading rate was set at 2 g/(L d) and the solid retention time at 40 days. When wheat straw alone was used as the substrate, the methane yields achieved with digestate liquor recycling was on average 240 ml CH₄/g VS giving a 21% improvement over the processes without recycling. However, over time, the processes suffered from declining methane yields and poor stability evidenced by low pH. To maintain process stability, wheat straw was co-digested with sewage sludge or supplemented with macronutrients (nitrogen and phosphorous). As a result, the processes with digestate liquor recycling could be operated stably, achieving methane yields ranging from 288 to 296 ml CH₄/g VS. Besides, the processes could not be operated sturdily with supplementation of macronutrients without digestate liquor recycling. The highest methane yield (296 ± 16 ml CH₄/g VS) was achieved by co-digestion with sewage sludge plus recycling of digestate liquor after filtration (retention of nutrients and microorganisms). This was comparable to the maximum expected methane yield of 293 ± 13 ml CH₄/g VS achieved in batch test. The present study therefore demonstrated that digestate liquor recycling could lead to a decreased dilution of vital nutrients from the reactors thereby rendering high process performance and stability.     

Sunflower as a biofuels crop: An analysis of lignocellulosic chemical properties

Four accessions of cultivated sunflower (Helianthus annuus) and silverleaf sunflower (Helianthus argophyllus), were each grown in three locations (Georgia, British Columbia, and Iowa) at different planting densities and phenotyped for biomass-related traits and wood biochemistry. In most environments, H. argophyllus produced significantly more biomass than H. annuus. Cell wall chemistry for a subset of plants grown in Georgia and Iowa was assessed using analytical wet chemistry methods to measure lignin and sugar content/composition. The analysis of lignin and the S/G-lignin ratios for a larger number of samples (n > 250) was also assessed by high-throughput pyrolysis Molecular Beam Mass Spectrometry. Average pyMBMS estimated lignin content (i.e., dry weight fraction) for 60 °C dried basal stem samples of H. annuus and H. argophyllus was 29.6% (range, 24.0%–34.6%) and 28.6% (range, 24.6%–33.3%), respectively when averaged across all environments. The average S/G lignin mass ratio was 1.5 (range, 1.0–2.0) for H. annuus and 1.7 (range, 1.0–2.4) in H. argophyllus. Stem samples from these two species only differed statistically for a few cell wall chemistry traits; however, accession level differences within each species were apparent. Cell wall chemistry in both species was significantly affected by both location and planting density, thus demonstrating the need to select for these traits in the environment for which the crop will be produced. Overall, these results show that cultivated sunflower and silverleaf sunflower both possess the necessary phenotypic diversity to facilitate the development of a hybrid sunflower with improved lignocellulosic biofuels traits, namely increased biomass, decreased lignin, and increased glucan.     

Second-generation bioethanol of hydrothermally pretreated stover biomass from maize genotypes

Twelve maize genotypes, were agronomically evaluated and their stover hydrothermally pretreated in a temperature range of 210–225 °C to assess the effects of genotype and pretreatment severity on stover recalcitrance toward bioethanol conversion. Maize genotypes exhibited significant variation for biomass yield and all agronomic evaluated, while among all cell wall constituents measured in the unpretreated stover, only ash content showed differences among genotypes. The pretreatment severities assayed impacted most stover compositional traits, and the glucose recovered after enzymatic hydrolysis displayed a similar profile among genotypes with similar genetic background. Harsher pretreatment conditions maximized the potential cellulosic bioethanol production (208–239 L/t), while the mildest maximized the bioethanol from the hemicellulosic hydrolysates (137–175 L/t). Consequently, when both pentose and hexose sugars were considered, the total potential bioethanol produced at the lowest and highest pretreatment temperatures was similar in all genotypes (292–358 L/t), indicating that the lowest temperature (210 °C) was the optimal among all assayed. Importantly, the ranking of genotypes for bioethanol yield (L/ha) closely resembled the ranking for stover yield (t/ha), indicating that breeding for biomass yield would increase the bioethanol production per hectare regardless of the manufacturing process. Similarly, the genetic regulation of corn stover moisture is possible and relevant for efficient energy production as biomass moisture has a potential impact on stover transportation, storage and processing requirements. Overall, these results indicate that local landrace populations are important genetic resources to improve cultivated crops, and that simultaneous breeding for production of grain and stover bioethanol is possible in corn.     

The improved CO2 capture system with heat recovery based on absorption heat transformer and flash evaporator

Absorption heat transformer (AHT) and flash evaporator (FE) are used to reduce the heat consumption of CO₂ capture processes and an AHT–FE-aided capture system is proposed. Analyses are carried out to verify the effectiveness in reducing heat consumption. Compared with the base CO₂ capture system of 3000 t/d CO₂ capture capacity from a 660 MW coal-fired power unit, the AHT–FE-aided capture system reduces the heat consumption from 3.873 GJ/tCO₂ to 3.772 GJ/tCO₂, and the corresponding energy saving is 2.62%. The economic analysis shows that the annual profit would be 2.94 million RMB Yuan. The payback period of the AHT–FE-aided capture system is approximately 2.4 years. Therefore, the AHT–FE-aided capture system is both economically and technically feasible for improving the CO₂ capture energy performance.     

Relationships between cellulosic biomass particle size and enzymatic hydrolysis sugar yield: Analysis of inconsistent reports in the literature

Cellulosic ethanol made from cellulosic biomass is a promising alternative to petroleum-based transportation fuels. Enzymatic hydrolysis is a crucial step in cellulosic ethanol production. In order to better understand the mechanisms of enzymatic hydrolysis, relationships between cellulosic biomass particle size and enzymatic hydrolysis sugar yield have been studied extensively. However, the literature contains inconsistent reports. This paper presents an analysis of the inconsistent reports on the relationships in the literature. It discusses the differences in the reported experiments from five perspectives (biomass category, particle size definition, sugar yield definition, biomass treatment procedure, and particle size level). It also proposes future research activities that can provide further understanding of the relationships.     

GIS-based assessment of sustainable crop residue potentials in European regions

In this paper a novel model based on a geographic information system (GIS) is presented for the assessment of sustainable crop residue potentials. The approach is applied to analyse the amount and the spatial distribution (1 km × 1 km grid cells) of cereal straw, root crop and oil plant residues for five European regions, considering spatially differentiated environmental sustainability issues, i.e. organic carbon content in topsoil, soil erodibility, and protected areas. The maximum sustainable residue potential varies strongly between the regions and residue types. In the scenarios Basis and Restrict, it accounts for 45–59% and 24–48% of the theoretical potential respectively without considering competing uses. Among the crop residues, cereal straw shows the highest energy potential in all regions under investigation. In terms of wet mass it accounts for 3.7 Mio. t_wet/a in North Rhine-Westphalia, 1.6 Mio. t_wet/a in Île-the-France, 1.2 Mio. t_wet/a in Wallonia, 0.9 Mio. t_wet/a in West Midlands, and 0.3 Mio. t_wet/a in South Netherlands (scenario Basis). Our survey shows that spatially differentiated potential estimations and the inclusion of crop residues other than cereal straw are urgently needed to improve the present rough estimations for crop residues which can be used in a sustainable way. The rather high spatial resolution of our analyses particularly allows for the support of regional stakeholders and prospective investors when it comes to questions of regional availability of biomass resources, transport distances to biomass conversion plants, and identification of suitable plant sites and sizes, respectively.     

Supercritical water gasification of timothy grass as an energy crop in the presence of alkali carbonate and hydroxide catalysts

This study is focused on identifying the candidature of timothy grass as an energy crop for hydrogen-rich syngas production through supercritical water gasification. Timothy grass was gasified in supercritical water to investigate the impacts of temperature (450–650 °C), biomass-to-water ratio (1:4 and 1:8) and reaction time (15–45 min) in the pressure range of 23–25 MPa. The impacts of carbonate catalysts (e.g., Na₂CO₃ and K₂CO₃) and hydroxide catalysts (e.g., NaOH and KOH) at variable mass fractions (1–3%) were examined to maximize hydrogen yields. In the non-catalytic gasification of timothy grass, highest hydrogen (5.15 mol kg⁻¹) and total gas yields (17.2 mol kg⁻¹) with greater carbon gasification efficiency (33%) and lower heating value (2.21 MJ m⁻³) of the gas products were obtained at 650 °C with 1:8 biomass-to-water ratio for 45 min. However, KOH at 3% mass fraction maximized hydrogen and total gas yields up to 8.91 and 30.6 mol kg⁻¹, respectively. Nevertheless, NaOH demonstrated highest carbon gasification efficiency (61.3%) and enhanced lower heating value of the gas products (4.68 MJ m⁻³). Timothy grass biochars were characterized through Fourier transform infrared spectroscopy, Raman spectroscopy and scanning electron microscopy to understand the behavior of the feedstock to rising temperature and reaction time. The overall findings suggest that timothy grass is a promising feedstock for hydrogen production via supercritical water gasification.     

Continuous liquid-phase valorization of bio-ethanol towards bio-butanol over metal modified alumina

Commercial mixed-phase aluminum oxide was used as a heterogeneous catalyst support, providing slightly basic properties which are well-suited for the condensation of bio-ethanol to C₄ hydrocarbons, such as 1-butanol. Different metals (Cu, Ni and Co), at various metal loadings were deposited on the support. Consequently, the catalytic reactions were carried out in a continuous laboratory-scale fixed bed reactor operated at 240 °C and 70 bar. The catalysts were characterized by means of XRD, TEM, FT-IR, XPS and ICP-OES. Different metals were found to give entirely different product distributions. With the best catalysts, the selectivities towards 1-butanol close to 70% were reached, while the ethanol conversion typically varied between 10 and 30% – strongly depending on the metal applied. It was observed that low loading of copper and high loading of nickel were responsible for the formation of 1-butanol, whereas cobalt and high loading of copper resulted in the production of ethyl acetate. The reaction was found to be extremely sensitive to catalyst preparation conditions and procedures such as metal loading, calcination/reduction temperature and, thereby, to the formation of corresponding crystallite structure.     

Assessing sustainable forest biomass potential and bioenergy implications for the northern Lake States region,USA

Forestlands in the United States have tremendous potential for providing feedstocks necessary to meet emerging renewable energy standards. The Lake States region is one area recognized for its high potential of supplying forest-derived biomass; however, the long-term availability of roundwood harvests and associated residues from this region has not been fully explored. Better distribution and temporal availability estimates are needed to formulate emerging state policies regarding renewable energy development. We used a novel predictive methodology to quantify sustainable biomass availability and likely harvest levels over a 100-year period in the Lake States region. USDA Forest Inventory and Analysis estimates of timberland were combined with published growth and yield models, and historic harvest data using the Forest Age Class Change Simulator (FACCS) to generate availability estimates. Monte-Carlo simulation was used to develop probability distributions of biomass harvests and to incorporate the uncertainty of future harvest levels. Our results indicate that 11.27–15.71 Mt y⁻¹ dry roundwood could be sustainably harvested from the Lake States. Assuming 65% collection rate, 1.87–2.62 Mt y⁻¹ residue could be removed, which if substituted for coal would generate 2.12–2.99 GW h of electricity on equivalent energy basis while reducing GHG (CO₂e) emission by 1.91–2.69 Mt annually. In addition to promoting energy security and reducing GHG emissions, forest residues for energy may create additional revenues and employment opportunities in a region historically dependent on forest-based industries.     

Lignocellulosic ethanol production employing immobilized Saccharomyces cerevisiae in packed bed reactor

Most of ethanol production processes are limited by lower ethanol production rate and recyclability problem of ethanologenic organism. In the present study, immobilized co-fermenting Saccharomyces cerevisiae GSE1618 was employed for ethanol fermentation using rice straw enzymatic hydrolysate in a packed bed reactor (PBR). The immobilization of S. cerevisiae was performed by entrapment in Ca-alginate for optimization of ethanol production by varying alginic acid concentration, bead size, glucose concentration, temperature and hardening time. Remarkably, extra hardened beads (EHB) immobilized with S. cerevisiae could be used up to repeated 40 fermentation batches. In continuous PBR, maximum 81.82 g L⁻¹ ethanol was obtained with 29.95 g L⁻¹ h⁻¹ productivity with initial glucose concentration of 180 g L⁻¹ in feed at dilution rate of 0.37 h⁻¹. However, maximum ethanol concentration of 40.33 g L⁻¹ (99% yield) with 24.61 g L⁻¹ h⁻¹ productivity was attained at 0.61 h⁻¹ dilution rate in fermentation of un-detoxified rice straw enzymatic hydrolysate (REH). At commercial scale, EHB has great potential for continuous ethanol production with high productivity using lignocellulosic hydrolysate in PBR.