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This study investigates a fed-batch simultaneous saccharification fermentation (F-SSF) process to increase ethanol titres from highly lignified (41.6 wt.%) cotton gin trash residue. The optimal initial solid loading, enzyme dose, feed quantities and intervals to maximize substrate feed and subsequent ethanol titres were examined. Under batch SSF conditions, initial extracted cotton gin trash (ECGT) solid loadings were maximised at 19.35 wt.% and attained an ethanol titre of 23.3 g/l with a corresponding yield of 53.7%. Operating under optimised F-SSF mode, fermentations were initiated with 16.13 wt% EGCT solids followed by fresh ECGT feeds of 16.13 wt% and 12.9 wt.% at 12-h intervals. Cellulase levels were maintained at 44 FPU/g glucan throughout the fermentations. The final ethanol titre of 41 .4 g/l with a corresponding conversion rate of 70.1% was achieved after 72 h. Comparable ethanol yields of 40 g/l with 67.8% conversion were realized with lower cellulase dosing (25 FPU g/glucan) but only after extending the fermentation by 24 h.

This study reports on the evaluation of a two-stage pretreatment process for preparing cotton gin trash (CGT) for conversation to ethanol. During the first stage, CGT was mixed with acid at 12% H2SO4 on solids and heated to 180 °C for 15 min in a pressurised stirred reactor. Pressed first-stage pretreated fibres were heated to 200 °C for 5 min during the second stage. The two-stage process facilitated excellent sugar recovery from both cellulose (84%) and hemicellulose (78%) fractions of CGT. Recombinant yeast GSF335 propagated on first-stage liquors yielded 41.8 g kg−1 of dry CGT and was compared with the commercial yeast during separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentations (SSF). SHF of pretreated CGT fibre yielded the equivalent of 66.2 and 64.0 kg ethanol per tonne of unprocessed CGT with GSF335 and the commercial yeast, respectively. SSF ethanol yields for the commercial yeast were significantly lower (50.3 kg) while GSF335 correspondingly produced 63.8 kg ethanol per tonne of raw CGT.

We evaluated a two stage pretreatment process for liberation of sugars from cotton gin trash (CGT) for ethanol production. The 1st stage releases C5 sugars for propagation of yeast inoculum intended for C6 sugar fermentations. The effects of a range of dilute acid hydrolysis pretreatment conditions on CGT were assessed. Propagation of recombinant Saccharomyces cerevisiae GSF335 in 1st stage liquors yielded up to 66 g yeast biomass kg−1 CGT. In the presence of PEG 6000, 2nd stage pretreated fibres were highly acquiescent to enzyme hydrolysis, yielding up to 242 g glucose kg−1 CGT. We conclude that the optimum conditions for recovery of sugars from CGT are: 1:6 solids to liquids ratio with 12% H2SO4 wt. on solids at 180 °C for 15 min during the 1st stage, with the ensuing pressed unwashed fibre heated to 200 °C for 5 min during the 2nd stage.

Soil degradation is a multi-factorial process that threatens crop production and the environment. According to the classification method of Pieri et al. (1992), soils are classified into different degradation classes by calculating the soil structural stability index (SSI). Soil nitrogen, phosphorus, labile organic carbon, mineral N and available-P concentration and stoichiometry under different degrees of degradation in rice fields in China were determined and analyzed. The results showed that 47% of the soil in China's rice fields had degradation problems. As the soil gradually degraded, soil N, P, AP concentrations were found to significantly decline (p

© 2017 Elsevier Ltd Optimisation of conditions for gamma-valerolactone (GVL) pretreatment of Australian eucalyptus sawdust for high cellulose biomass and bioethanol production was demonstrated. Pretreatment parameters investigated included GVL concentrations of 35-50% w/w, temperatures of 120-180 °C and reaction durations of 0.5-2.0 h. Optimum conditions were determined using the response surface method (RSM) and central composite face-centred design. Cellulose content increased from 39.9% to a maximum of 89.3% w/w using treatments with 50% GVL at 156 °C for 0.5 h. Temperature had the most significant effect (RSM p < .05) on cellulose content of residual biomass and reducing operational duration of < 0.5 h may be viable according to RSM. PSSF fermentations of optimised pretreated eucalyptus sawdust produced up to 94% theoretical ethanol yield, which corresponded to approximately 181 kg of ethanol per dry ton of eucalyptus sawdust. The compositions of both the residual biomass and pretreatment liquors show that GVL pretreatment is a promising solvent for lignocellulosic biorefining.

A two-stage dilute acid and steam explosion (SE) pretreatment process was developed and evaluated at pilot scale for ethanol production from cotton gin trash (CGT). Optimal conditions for CGT processing were defined as 1:6 solids to liquids ratio with 9% H2SO4 wt. on solids at 180 °C for 15 min. during stage 1 with ensuing pressed fibres successively exposed to SE at 200 °C for 5 min during stage 2. SE fibres were highly acquiescent to enzyme hydrolysis (76%) in the presence of PEG 6000, yielding 381 g glucose kg−1 fibre. Simultaneous saccharification and fermentation (SSF) trials validated the selected process option and additional fed-batch SSFs confirmed titres above the minimum 4% ww-1 benchmark for economically viable distillation. The practicality of converting CGT to ethanol was demonstrated at pilot scale with titres above 4% ww-1 and a conversion efficiency of 60% t−1 dry GCT.

Whole Eucalyptus grandis trees (including bark, branches and leaves) were investigated as a potential feedstock for bioethanol production. To demonstrate and maximize ethanol production, unwashed steam exploded wood chips (SEWC) were used as substrates in high solids load simultaneous saccharification fermentations (SSF) with an industrial Saccharomyces cerevisiae strain (Fali®). Under optimized SSF conditions—20 wt% solids, 60 filter paper units (FPU)/g glucan, and 36°C—ethanol titer and glucan-to-ethanol yields of 56 g/L and 90%, respectively, were achieved. Raising the SSF temperature to 39°C showed no observable benefit in final ethanol titers and/or yields. Addition of 3 wt% polyethylene glycol (PEG) further increased final ethanol titers and yields to 60 g/L and 95%, respectively, at a 30% lower cellulase dosage. Considering the mass balance of the best-performing SSF (20 wt% SEWC, 40 FPU Ctec 2/g glucan, 30 mg PEG/g dry solid at 36°C) process, the maximum ethanol attainable was estimated at 187 kg (85.8% yield) and 299 kg (95% yield) per dry metric ton of original and SEWC biomass, respectively.

This paper reports on mild acid pretreatment options for the conversion of Agave tequilana leaves into composite sugars for ethanol fermentation. The effect of five different pretreatment conditions (time, temperature and acid concentrations) were assessed in terms of cellulose digestibility, hemicellulose solubilisation and lignin content in leaves of 1.25 years old A. tequilana plants from Rockhampton and 2.5 year plants from Kalamia. Dilute acid pretreatment and enzyme saccharification of A. tequilana leaf bagasse significantly improved total glucose recovery. A recovery of 273 mg/g (70% theoretical) was attained when the bagasse was pretreated with 2.0% H2SO4 for 60 min at 121 °C and saccharified with 6% (w/w) CTec 2. Saccharomyces cerevisiae efficiently fermented crude A. tequilana bagasse and juice hydrolysates within 13 h and 7 h respectively, yielding up to 38.6 g/L and 12.4 g/L. This corresponds to glucose to ethanol conversion rate of 68 and 61% for A. tequilana leaf bagasse and juice, respectively. With further developments, including fermentation of C5 sugars and inulinase saccharification of juices (release of fructose), this process could deliver greater yields, reinforcing its potential as a biofuel feedstock.

The widespread deployment of commercial-scale cellulosic ethanol currently hinges on developing and evaluating scalable processes whilst broadening feedstock options. This study investigates whole Eucalyptus grandis trees as a potential feedstock and demonstrates dilute acid pre-treatment (with steam explosion) followed by pre-saccharification simultaneous saccharification fermentation process (PSSF) as a suitable, scalable strategy for the production of bioethanol. Biomass was pre-treated in dilute H2SO4 at laboratory scale (0.1 kg) and pilot scale (10 kg) to evaluate the effect of combined severity factor (CSF) on pre-treatment effectiveness. Subsequently, pilot-scale pre-treated residues (15 wt.%) were converted to ethanol in a PSSF process at 2 L and 300 L scales. Good polynomial correlations (n = 2) of CSF with hemicellulose removal and glucan digestibility with a minimum R2 of 0.91 were recorded. The laboratory-scale 72 h glucan digestibility and glucose yield was 68.0% and 51.3%, respectively, from biomass pre-treated at 190 °C /15 min/ 4.8 wt.% H2SO4. Pilot-scale pre-treatment (180 °C/ 15 min/2.4 wt.% H2SO4 followed by steam explosion) delivered higher glucan digestibility (71.8%) and glucose yield (63.6%). However, the ethanol yields using PSSF were calculated at 82.5 and 113 kg/ton of dry biomass for the pilot and the laboratory scales, respectively. © 2016 Society of Chemical Industry and John Wiley & Sons, Ltd

This paper reports on processing options for the conversion of feedlot cattle manures into composite sugars for ethanol fermentation. Small-scale anaerobic digestion trials revealed that the process significantly reduces the content of glucan and xylan (ca. 70%) without effecting lignin. Moreover, anaerobic digestate (AD) fibres were poor substrates for cellulase (Cellic® CTec 2) saccharification, generating a maximum combined sugar yield of ca. 12% per original dry weight. Dilute acid pretreatment and enzyme saccharification of raw manures significantly improved total sugar recoveries, totalling 264 mg/g (79% theoretical). This was attained when manures were pretreated with 2.5% H 2 SO 4 for 90 min at 121 °C and saccharified with 50 FPU CTec 2/g glucan. Saccharomyces cerevisiae efficiently fermented crude hydrolysates within 6 h, yielding 7.3 g/L ethanol, representing glucose to ethanol conversion rate of 70%. With further developments (i.e., fermentation of xylose), this process could deliver greater yields, reinforcing its potential as a biofuel feedstock.

Cotton gin trash (CGT), a waste product of cotton gins, make up about 10% of each bale of cotton bolls ginned. The current study investigates high value volatile compounds in CGT to add value to this by-product. The volatile compounds in CGT and different parts of the cotton plant were extracted using various methods, identified by gas chromatography-mass spectrometry (GC-MS) or nuclear magnetic resonance (NMR) spectroscopy, and then quantified by gas chromatography-flame ionisation detection (GC-FID) against available standards. Terpenoids including monoterpenoids and sesquiterpenoids were found to be the most abundant, making up 64.66% (area under peak) of total volatiles extracted by hydro-distillation. The major extractable terpenoids in CGT were α-pinene (13.69-23.05 μg/g), β-caryophyllene (3.99-74.32 μg/g), α-humulene (2.00-25.71 μg/g), caryophyllene oxide (41.50-102.08 μg/g) and β-bisabolol (40.05-137.32 μg/g). Recoveries varied between different extraction methods. The terpenoids were found to be more abundant in the calyx (659.12 μg/g) and leaves (627.72 μg/g) than in stalks (112.97 μg/g) and stems (24.24 μg/g) of the cotton plant, indicating the possible biological origin of CGT volatiles. This study is the first to identify and quantify the different terpenoids present in CGT and significantly, β-bisabolol, an abundant compound (sesquiterpene alcohol) which may have valuable biological prospects. These findings therefore contribute to identifying alternative management strategies and uses of CGT.

The effectiveness of high temperature, dilute H2SO4 and NaOH pretreatments for the conversion of a post extracted cotton gin trash (ECGT) biomass was investigated. Whole acidified ECGT slurries were pretreated at various temperatures (180–230 °C) with or without the addition of extra catalyst. Compositional analysis of recovered ECGT fibre revealed xylan removal increased with pretreatment temperature under low pH conditions. Alkaline pretreatment (20 wt% NaOH) selectively delignified the ECGT fibre by as much as 80%. Characterisation of structural features identified an increase in cellulose crystallinity and surface area for all pretreated fibres and positively correlated to increasing pretreatment severity. Pretreatment of acidified ECGT slurries at 230 °C resulted in the highest glucan digestibility (66%) and glucose yields (51%). Glucan digestibility showed a strong positive correlation to xylan removal and surface area but weaker to delignification and total crystallinity for all dilute acid pretreated fibres.

Lignocellulosic materials are potentially a relatively inexpensive and abundant feedstock for biofuel production. The key to unlocking lignocelluloses recalcitrance is in part, the development of an effective pretreatment process. A promising new pretreatment method for lignocellulosic materials is the use of ionic liquids (ILs). ILs are non-volatile solvents that exhibit unique solvating properties. In this review, the dissolution of cellulose and lignocellulose in various ionic liquids is described including key properties such as high hydrogen-bonding basicity, which increases the ability of the IL to dissolve cellulose. As a pretreatment in biofuel production, the review details aspects such as the regeneration of cellulose from ionic liquids, structural changes that arise in the regenerated cellulose and its effect on enzymatic hydrolysis, the potential for IL recycling and finally, exploiting ILs in an integrated bioprocess.

Conditions for optimal pretreatment of eucalypt ( Eucalyptus dunnii ) and spotted gum ( Corymbia citriodora ) forestry thinning residues for bioethanol production were empirically determined using a 3 3 factorial design. Up to 161 mg/g xylose (93% theoretical) was achieved at moderate combined severity factors (CSF) of 1.0–1.6. At CSF > 2.0, xylose levels declined, owing to degradation. Moreover at high CSF, depolymerisation of cellulose was evident and corresponded to glucose (155 mg/g, ∼33% cellulose) recovery in prehydrolysate. Likewise, efficient saccharification with Cellic® CTec 2 cellulase correlated well with increasing process severity. The best condition yielded 74% of the theoretical conversion and was attained at the height of severity (CSF of 2.48). Saccharomyces cerevisiae efficiently fermented crude E. dunnii hydrolysate within 30 h, yielding 18 g/L ethanol, representing a glucose to ethanol conversion rate of 0.475 g/g (92%). Based on our findings, eucalyptus forest thinnings represent a potential feedstock option for the emerging Australian biofuel industry.

Dilute sulphuric acid pretreatment followed by enzyme saccharification of Sorghum bicolor straw was undertaken to examine its potential as a feedstock in bioethanol production in Australia. Factorial design experiments evaluated the impact of pretreatment parameters on hemicellulose solubilisation and cellulose enzymatic hydrolysis. Sugar yields in prehydrolysate and saccharified liquors were found to increase with treatment severity; temperature was found to have the greatest impact. Degradation products were minimal; acetate and total phenolics peaked at 33 and 1.5 mg/g respectively. Conditions for maximum hemicellulose solubilisation (2% H 2 SO 4 for 60 min at 121 °C) differed to those associated with maximum glucose release from solid residue saccharifications (1% H 2 SO 4 /90 min /121 °C). Water extractive sugars accounted for over 20% total sugars recovered. Addition of β-glucosidase and xylanase to enzyme saccharification enhanced reaction rates and final sugar yields three-fold, whilst reducing cellulase dosage. Considering its abundance, high sugar potential and apparent ease of conversion, sorghum straw is an appropriate feedstock for the production of second generation fuels.

A porous iron-based oxalate-phosphate-amine metal-organic framework material (OPA-MOF) was investigated as a microbially-induced slow-release nitrogen (N) and phosphorus (P) fertilizer. Seedling growth, grain yields, nutrient uptake of wheat plants, and soil dynamics in incubated soil, were investigated using OPA-MOF vs standard P (triple-superphosphate) and N (urea) fertilizers in an acidic Ferralsol at two application rates (equivalent 120 and 40 kg N ha(-1)). While urea hydrolysis in the OPA-MOF treatment was rapid, conversion of ammonium to nitrate was significantly inhibited compared to urea treatment. Reduced wheat growth in OPA-MOF treatments was not caused by N-deficiency, but by limited P-bioavailability. Two likely reasons were slow P-mobilisation from the OPA-MOF or rapid P-binding in the acid soil. P-uptake and yield in OPA-MOF treatments were significantly higher than in nil-P controls, but significantly lower than in conventionally-fertilised plants. OPA-MOF showed potential as enhanced efficiency N fertilizer. However, as P-bioavailability was insufficient to meet plant demands, further work should determine if P-availability may be enhanced in alkaline soils, or whether central ions other than Fe, forming the inorganic metal-P framework in the MOF, may act as a more effective P-source in acid soils.

Physico-chemical pretreatment of lignocellulosic biomass is critical in removing substrate-specific barriers to cellulolytic enzyme attack. Alkaline pretreatment successfully delignifies biomass by disrupting the ester bonds cross-linking lignin and xylan, resulting in cellulose and hemicellulose enriched fractions. Here we report the use of dilute alkaline (NaOH) pretreatment followed by enzyme saccharifications of wheat straw to produce fermentable sugars. Specifically, we have assessed the impacts of varying pretreatment parameters (temperature, time and alkalinity) on enzymatic digestion of residual solid materials. Following pretreatment, recoverable solids and lignin contents were found to be inversely proportional to the severity of the pretreatment process. Elevating temperature and alkaline strengths maximised hemicellulose and lignin solubilisation and enhanced enzymatic saccharifications. Pretreating wheat straw with 2% NaOH for 30 min at 121 °C improved enzyme saccharification 6.3-fold when compared to control samples. Similarly, a 4.9-fold increase in total sugar yields from samples treated with 2% NaOH at 60 °C for 90min, confirmed the importance of alkali inclusion. A combination of three commercial enzyme preparations (cellulase, β-glucosidase and xylanase) was found to maximise monomeric sugar release, particularly for substrates with higher xylan contents. In essence, the combined enzyme activities increased total sugar release 1.65-fold and effectively reduced cellulase enzyme loadings 3-fold. Prehydrolysate liquors contained 4-fold more total phenolics compared to enzyme saccharification mixtures. Harsher pretreatment conditions provide saccharified hydrolysates with reduced phenolic content and greater fermentation potential.

In this study, Bauxsol pellets packed in PVC columns were used to remove nutrients and trace-metals from municipal wastewater during a 6 months field trial. Bauxsol pellet columns showed a high phosphate removal rate via precipitation of PO(4)(3-) with Ca(2+) and Mg(2+) ions: at 90% in the 1st month; at 80% from the second to fifth months; and at 60% in the sixth month. Pellet bound total phosphorus and Colwell phosphorus were 7.3 g/kg and 2 g/kg and are about 20 times the concentrations found in most fertile soils. Trace-metals in effluents were bound, probably irreversibly under the columns' environmental conditions, to the Bauxsol minerals that have high surface area to volume ratios and high charge to mass ratios. Experimental results showed a complex nitrogen cycle operating within the Bauxsol pellet columns including anoxic nitrification, denitrification, and anammox processes. Although a transient pH spike, associated with the release of unreacted CaO from the cement binder used in the pellets, was observed, this may be readily corrected through post-treatment pH adjustment. Hence, the geochemistry of Bauxsol pellets can effectively remove and bind nutrients and trace-metals during wastewater treatment, and further research may show that saturated spent pellets can be used as fertilizer.

Mild alkali cooking of lignocellulosic biomass is an effective pretreatment method, which improves enzymatic hydrolysis. Here, we report the use of dilute alkali (NaOH) pretreatment followed by enzyme saccharification of cereal residues for their potential to serve as feedstock in the production of next-generation biofuels in Australia. After pretreatment, both solids and lignin content were found to be inversely proportional to treatment severity. We also found that higher temperatures and alkali strength were quintessential for maximizing sugar recoveries from enzyme saccharifications. Generally, pretreatment conditions at elevated temperatures led to highly digestible material enriched in both cellulose and hemicellulose components. Increasing cellulase loadings and tailoring enzyme activities with additional β-glucosidases and xylanases delivered greater rates of monosaccharide sugar release and yields throughout enzyme hydrolysis. Considering their abundance, high sugar potential, and apparent ease o...

BACKGROUND: Prehydrolysis of wheat stubble using moderate temperatures and dilute acid strength is an effective means for solubilizing hemicellulose fractions and improving cellulose hydrolysis. Variation in prehydrolysis parameters (temperature, time, and acid strength) and enzymatic saccharification conditions were examined for conversion of wheat stubble into fermentable sugars. RESULTS: Elevating temperature and acid strength maximized sugar release in prehydrolysate liquors. The optimal conditions of 2.0% H2SO4/60 min/121 °C effectively solubilized 79% of the available hemicellulose. Production of inhibitory hydrolysis and degradation products such as acetic acid and levulinic acid, were detected at levels of 3.4 g L−1 and 0.64 g L−1, respectively. Sugar yields in prehydrolysate and saccharified liquors were found to increase with treatment severity. Temperature had the greatest impact on sugar release, followed by acid concentration and time. Optimizing prehydrolysis conditions at 1.0% H2SO4/90 min/121 °C, produced a 3.2-fold improvement in cellulose hydrolysis with recoveries approaching 82%. The addition of β-glucosidase and xylanase to the cellulase preparations assisted monomeric sugar release. CONCLUSION: Although treatment conditions for hemicellulose and cellulose hydrolysis differ, the study's findings suggest a good degree of overlap and process flexibility which should permit high recovery of pentose and hexose sugars. Copyright © 2011 Society of Chemical Industry

The impacts of varying pretreatment parameters (temperature, time, and alkalinity) on enzymatic hydrolysis of sorghum straw were investigated. Following pretreatment, both solids and lignin content was found to be inversely proportional to the severity of the treatments. Higher temperatures and alkali strength were quintessential for maximising sugar recoveries from enzyme saccharifications. Total sugar release peaked when sorghum straw was pretreated in 2% NaOH at 121 degrees C for 60 min; representing a 5.6-fold higher yield compared to samples pretreated at 60 degrees C in the absence of alkali. Similarly, 4.3-fold increases in total sugars from samples treated with 2% NaOH at 60 degrees C for 90 min, confirmed the importance of alkali inclusion. Addition of beta-glucosidase and xylanase to saccharification mixtures enhanced reaction rates and final sugar yields, whilst reducing cellulase dosage 4-fold. Saccharification efficiency of pretreated solids approached 90% and 95% (w/w) with as little as 2.5 and 5.0 FPU cellulase/g, respectively.

Bauxsol reagents (powder, slurry, or pellet forms) are powerful tools in environmental remediation and water and sewage treatment However, when used in circum-neutral water treatments, cement-bound Bauxsol pellets produce a sustained pH and alkalinity spike due to the presence of unreacted CaO in the cement binder. This study developed a pellet treatment system to minimize the alkalinity/pH spike. The recipe for pelletization consisted of Bauxsol powder, ordinary Portland cement (OPC), hydrophilic fumed silica, aluminum powder, a viscosity modifier, and water. Several batches (including different ratios and sizes) were run using modified makeup waters (H(2)0 + CO(2) or NaHCO(3)) or curing brines (CO(2), NaHCO(3), or Mg/CaCl(2)). Alkalinity, pH stability, and slake durability tests were performed on pellets before and/or after curing. The best result for reducing the alkalinity/pH spike was obtained from a MgCl(2), CaCl(2) bath treatment using a Bauxsol:cement ratio of 2.8:1 (pH 8.28; alkalinity 75.1 mg/L) for a 100 g batch or 245:1 (pH 8.05; alkalinity 35.4 mg/L) for a 1 kg batch. Although brine curing does provide a control on pH/alkalinity release, the pellets may still contain unreacted CaO. Therefore, a freshwater rinse of pellets before treating circum-neutral waters is recommended as is the continued investigation of alternative pellet binders.

In this study, dilute acid pretreatment of five biomass feedstocks viz., sugarcane trash, sugarcane bagasse, rice straw, corn stover and palm empty fruit bunch were compared at a given combined severity factor (CSF) range of 1.4–3.2 and were characterised using an alternative Simons’ staining dye—Direct Yellow 11 fraction (DY 11, molecular weight >100,000) to better understand the correlations of pretreatment effectiveness with biomass physicochemical properties and pretreatment conditions. Good polynomial correlations (n = 2) of CSF were obtained with hemicellulose removal, cellulose digestibility and glucose yield resulting in R2 > 0.95. The results show that the total contents of extractives and ash have negative impacts on dilute acid pretreatment. Simons’ staining results show that DY 11 can also be used to estimate cellulose accessibility to cellulase enzymes. Good linear correlations of maximum adsorption capacity of DY 11 with CSF (R2 = 0.87–0.99) and cellulose digestibility (R2 = 0.91–0.99) were observed for most of the pretreated biomass samples.

Cotton ginning trash (CGT) collected from Australian cotton gins was evaluated for bioethanol production. CGT composition varied between ginning operations and contained high levels of extractives (26-28%), acid-insoluble material (17-22%) and holocellulose (42-50%). Pretreatment conditions of time (4-20 min), temperature (160-220 °C) and sulfuric acid concentration (0-2%) were optimised using a central composite design. Response surface modelling revealed that CGT fibre pretreated at 180 °C in 0.8% H2SO4 for 12 min was optimal for maximising enzymatic glucose recoveries and achieved yields of 89% theoretical, whilst the total accumulated levels of furans and acetic acid remained relatively low at1 and 2 g/L respectively. Response surface modelling also estimated maximum xylose recovery in pretreated liquors (87% theoretical) under the set conditions of 150 °C in 1.9% H2SO4 for 23.8 min. Yeast fermentations yielded high ethanol titres of 85%, 88% and 70% theoretical from glucose generated from: (a) enzymatic hydrolysis of washed pretreated fibres, (b) enzymatic hydrolysis of whole pretreated slurries and (c) simultaneous saccharification fermentations, respectively.

Municipal wastewater was treated over a six month period in an unplanted constructed wetland with a lower soil layer and an upper Bauxsol™ pellet layer. The interactions between Bauxsol™ pellets, soil, effluent and microbial communities demonstrated a positive influence on contaminant removal. Bauxsol™ treated effluent showed > 95% phosphate removal and ~ 26% nitrogen removal during the trial. Substantial quantities of nitrate, trace-metals and Colwell P were bound to the pellets, whereas only ammonium was bound to the soil. The structure of microbial communities analysed by denaturing gradient gel electrophoresis (DGGE) showed distinct bacterial communities attached to Bauxsol™ pellets and soil owing to differences in geochemistry and micro-environmental conditions. Polymerase chain reaction (PCR) amplification of specific marker genes (i.e. bacterial and archaeal amoA genes, nosZ gene, and hzo gene) was used to evaluate the presence of microbial communities associated with nitrogen transformation. Data revealed the co-existence of aerobic ammonia-oxidising bacteria, anaerobic ammonia-oxidising bacteria (anammox) and denitrifiers attached to Bauxsol™ pellets and ammonia-oxidising bacteria and archaea attached to soil. This study successfully demonstrates that Bauxsol™ pellets are a suited alternative media for constructed wetland to treat wastewater effectively removing phosphate and serving as biomass support particles for bacterial communities associated with nitrogen-cycling.

Promoter libraries of Zymomonas mobilis were generated in Escherichia coli using the CAT promoterless vector, pKK232-8. Among the 15 clones retained for further study, quantitative analysis of promoter strength revealed a 140-fold difference between the weakest and strongest promoter fragment. Nucleotide sequence determination and primer extension analysis were used to locate possible promoter-like sequences. Inspection of the DNA sequence surrounding the mRNA start-site, revealed regions similar to the E. coli sigma 70 consensus sequence (TTGACA-17 bpsTATAAT), approximately 4-8 bps upstream. Analogous to E. coli upstream activators, regions high in AT content (70-80%) and rich in static DNA bands, were found preceding the − 35 hexamer. Only one promoter fragment clone (Z1) was found to carry an open reading frame preceded by a sequence resembling a ribosome-binding site with a free energy of SD-anti-SD binding (ΔG°) of − 12.4 kcal/mol

A new method for extracting soil enzymes is described and a microplate method for assaying soil β-1,4-glucanases (cellulases) and β-1,3-glucanases (laminarinases). Soil samples were mechanically disrupted to produce crude enzyme extracts, and diluted preps incubated in microplates containing either carboxymethyl cellulose (CMC) to determine cellulase activity or laminarin substrate to determine laminarinase activity. The resulting glucose was measured using the fluorometric Amplex Red ® glucose assay. The method was reproducible, could be completed in 1 day and measured twice as much enzyme activity than the standard passive soil enzyme extraction procedure. The method described herein facilitates the development of high-throughput soil multiplex enzymatic assays from several soil samples at one time, and is well suited to the study of functional microbial ecology.

Aims To develop a method to produce beads with encapsulated Rhodococcus erythropolis NI86/21 with high cell density, extended shelf life, ease of handling and good atrazine degradation capabilities in both liquid and in agricultural soil. Methods and results Our findings show that the supplementary recovery step in nutrient broth media shortly after cell encapsulation facilitates cell survival in both wet and dry beads upon extended storage at 4 degrees C. Air drying has little or no impact on encapsulated R. erythropolis cell's ability to degrade atrazine in liquid or soil. Bead storage for periods extending up to 12 months at 4 degrees C did not affect the capacity of R. erythropolis encapsulated cells to degrade atrazine in either BMN or nonsterile soil extracts. Bentonite-amended beads formulated with 1% skim milk and exposed to the supplementary growth step, outperformed all other bead formats. These beads provided adequate numbers of vigorous R. erythropolis cells in either liquid or soil media to degrade atrazine. Conclusions Supplementary growth in nutrient broth media immediately following cell encapsulation greatly enhances R. erythropolis cells survival in both wet and dry beads upon extended storage at 4 degrees C. Wet and dried beads have similar capacity for atrazine degradation, and their usefulness and appeal in agronomic practise will only be known after bioassay evaluation and successful demonstration at field scale using incurred residues. Significance and impact of the study R. erythropolis NI86/21 encapsulated cells have the potential to reduce residual atrazine in soil, thereby minimizing the likelihood of off-site transport to ground or river water and reduce the loss of crops because of phytotoxicity of residual herbicide. Owing to their ease of handling, storage and possible compatibilities with pre-existing mechanical equipment, dried bead formats are ideally suited for agricultural and remediational applications.

Aims: To develop an encapsulation procedure for Rhodococcus erythropolis NI86/21 and demonstrate its use as a slow-release inoculant for reducing atrazine levels in aquatic and terrestrial environments. Methods and Results: Alginate encapsulation procedures were developed for the atrazine-degrading bacteria R. erythropolis NI86/21. Several bead amendments, including bentonite, powdered activated carbon (PAC) and skimmed milk (SM), were evaluated for slow release of R. erythropolis NI86/21 and efficacy of atrazine degradation. All bead types demonstrated a capacity to degrade atrazine in basal minimal nutrient buffer whilst continually releasing viable bacterial cells. We found that the addition of bentonite hastened cell release whilst SM sustained cell viability in bead formulations. Reducing the percentage of SM to 1% (w/v) resulted in faster rates of atrazine degradation in both liquid and soil, and was found to prolong cell survival upon bead storage. Limited oxygen transfer affects the capacity of the encapsulated R. erythropolis cells to degrade atrazine. Conclusions: Degradation studies have demonstrated the efficacy of R. erythropolis encapsulated cells to degrade atrazine in amended liquid and soil. However, in their current formulation, the wet alginate-based beads are impractical for field application because of their poor cell viability during storage. Significance and Impact of the Study: R. erythropolis NI86/21-encapsulated cells have the potential to reduce atrazine residues in a number of soil and water environments, possibly ensuring the continued registration and use of atrazine in agriculture by minimizing or eliminating nontarget effects.

The organophosphate pesticide, Ethion, remains a major environmental contaminant in rural Australia and poses a significant threat to environmental and public health. The aerobic degradation of Ethion by mesophilic bacteria isolated from contaminated soils surrounding disused cattle dip sites was investigated. Two isolates, identified as Pseudomonas and Azospirillum species, were capable of biodegrading Ethion when cultivated in minimal salts medium. The abiotic hydrolytic degradation products of Ethion such as Ethion Dioxon and O,O-diethylthiosphosphate were not detected. The data suggest the rapid degradation of Ethion to support microbial growth. The results have implications for the development of a bioremediation strategy.