Comparison between solid-state and powder-state alkali pretreatment on saccharification and fermentation for bioethanol production from rice straw

The efficacy of different concentrations of NaOH (0.25%, 0.50%, 0.75%, and 1.00%) for the pretreatment of rice straw in solid and powder state in enzymatic saccharification and fermentation for the production of bioethanol was evaluated. A greater amount of biomass was recovered through solid-state pretreatment (3.74 g) from 5 g of rice straw. The highest increase in the volume of rice straw powder as a result of swelling was observed with 1.00% NaOH pretreatment (48.07%), which was statistically identical to 0.75% NaOH pretreatment (32.31%). The surface of rice straw was disrupted by the 0.75% NaOH and 1.00% NaOH pretreated samples as observed using field-emission scanning electron microscopy (FE-SEM) and atomic force microscopy (AFM). In Fourier-transform infrared (FT-IR) spectra, absorbance of hydroxyl groups at 1,050 cm^?1 due to the OH group of lignin was gradually decreased with the increase of NaOH concentration. The greatest amounts of glucose and ethanol were obtained in 1.00% NaOH solid-state pretreated and powder-state hydrolyzed samples (0.804 g g^?1 and 0.379 g g^?1, respectively), which was statistically similar to the use of 0.75% NaOH (0.763 g g^?1 and 0.358 g g^?1, respectively). Thus, solid-state pretreatment with 0.75% NaOH and powder-state hydrolysis appear to be suitable for fermentation and bioethanol production from rice straw.     

The effect on digestibility of a new technique for alkali treatment of straw

The Biotechnical Institute in Kolding, Denmark, in collaboration with the National Institute of Animal Science, Copenhagen, has developed a new method for dry chemical treatment of straw with NaOH. Treatment with up to 7% NaOH (percent of straw dry matter) linearly increased enzyme digestibility and digestibility in vitro, and also rate of digestion. Lye treatment reduced cell wall constituents in straw (NDF). Digestibility in sheep was increased by increments of NaOH up to, but not exceeding, 4–5%. Although the process omits washing of excess base from the product, unreacted NaOH did not appear great enough to create concern.     

Alkaline treatment of wheat straw for increasing anaerobic biodegradability

Wheat straw was treated with NaOH and anaerobically digested for methane production. Alkaline treatment resulted in a greater than 100% increase in biodegradability of wheat straw. The potential of a process flow scheme employing high alkali concentration at ambient temperature with solids separation and recycle of filtrate containing residual alkali was explored. The effect of NaOH on the solubilization of cell wall constituents and potential problems of toxicity are discussed. A solubilization model was developed which is used to predict biodegradability of whole samples based on solids and filtrate biodegradabilities. Energy requirements and chemical costs are also addressed.     

Utilization of alkali-treated and neutralized wheat straw-based rations for growing goat kids

Six groups of six goat kids were fed individually for 168 days with wheat straw given various treatments: (1) control; (2) 33 g NaOH/kg straw; (3) 80 g NaOH/kg, partly neutralized with mineral acids; (4) mineral control for 80 g NaOH/kg; (5) 120 g NaOH/kg, partly neutralized with mineral acids, and (6) mineral control for 120 g NaOH/kg straw. The average weight gain was significantly superior (P< 0.05) and the efficiency of dry matter (DM) and energy utilization was the highest with the 80 g NaOH/kg straw treatment. This treatment also gave significantly higher (P<0.05) digestibility of DM, organic matter (OM), neutral detergent fibre (NDF), acid detergent fibre (ADF), nitrogen-free extract (NFE) and hemicellulose than the control and 33 g NaOH/kg straw treatments. Increasing levels of alkali decreased (P<0.05) the digestibility of crude protein (CP) and ether extract (EE). Digestible energy and nitrogen-corrected metabolisable energy (ME_n) (as a percentage of gross energy (GE)) were maximal with 80 g NaOH/kg. The pH value of rumen liquor was the same for the control and the 33 g NaOH/kg and 80 g NaOH/kg treatments, but significantly increased (P<0.05) with the 120 g NaOH/kg straw treatment. The mean values for rumen ammonia nitrogen (NH₃ -N) were the same for the control, the 33 g NaOH/kg, and mineral controls for 80 and 120 g NaOH/kg treatments, but 80 g NaOH and 120 g NaOH/kg straw gave significantly lower values. It is suggested that by partially neutralizing the residual alkali, 80 g NaOH/kg straw can give higher efficiency of energy utilization for growth and digestibility of nutrients compared with 33 g NaOH/kg or the untreated control group, and the extensive use of treated straw in the diets of animals of which a rapid rate of production is not demanded, may be advantageous.     

Ensiled alkali-treated straw. I. Effect of level and type of alkali on the composition and digestibility in vitro of ensiled barley straw

In three experiments barley straw chopped to 5 cm nominal particle length was ensiled in laboratory silos for 90 days after treatment with alkali. In the first two experiments, NaOH was added at 0, 1.05, 2.10, 3.15 or 4.20 g per 100 g straw dry matter (DM) (Experiment 1) or at 0, 2.5, 5.0, 7.5 and 10.0 g per 100 g straw DM (Experiment 2) in solutions at either 60 ml or 120 ml solution per 100 g straw DM. Digestibility in vitro of organic matter (OM) and digestible OM in DM (DOMD) increased with increasing level of NaOH. The effect of volume of solution on digestibility was small. The pH of the straws decreased during storage. The content of neutral detergent fibre decreased as the level of NaOH increased, but there was relatively little change in the contents of acid detergent fibre or acid detergent lignin. Lactate and acetate were detected in all silages, and butyrate was present in silages made from straws treated with less than 5 g NaOH per 100 g straw DM. On opening the silos little moulding was seen and the temperature of the straws remained close to ambient in both experiments throughout 16 days of subsequent exposure to air.In the third experiment, the comparative effects of Ca(OH)₂ and KOH were studied alone and in combination ($\text{50}\text{50}$ by weight) with NaOH. KOH mixed with NaOH gave levels of DOMD in vitro similar to those obtained with NaOH alone. Ca(OH)₂, whilst improving DOMD, was slightly less effective than the other alkalis.The optimum level of alkali for the treatment of barley straw prior to ensiling appeared to be 7.5 g/100 g straw DM. At this level of addition, DOMD in vitro would be expected to be about 65%. Ca(OH)₂ is worth further attention as an alternative to NaOH.     

Alkali treatment and fermentation of straw for animal feed

The feed value of annual ryegrass straw was improved by treatment with various concentrations of NaOH or NH₃ followed by fermentation of the treated straw with a mixed culture of Cellulomonas sp. and Alcaligenes faecalis. Laboratory feeding trials with voles showed that NaOH or NH₃ treatment considerably increased the feed efficiency of straw, but apparently gave a poorly palatable product. Fermentation tended to decrease the in vitro rumen digestibility (IVRD) of alkali-treated straw. The fermentations were carried out aerobically on a semisolid straw matrix having 11–86% moisture. Treatment by both NaOH and NH₃ increased the IVRD of straw. NH₃ also increased the nitrogen content in straw. The optimum condition for alkaline treatment of the straw was 4–6% NaOH for 1 hr or with 3% NH₃ for four weeks at room temperature. A minimum of 63% moisture was needed for significant fermentation of the straw. The combined effects of NaOH treatment and fermentation more than doubled crude protein, doubled crude fat, and increased IVRD by 75%. The NH₃ plus fermentation treatment tripled crude protein, doubled crude fat, and increased IVRD by 60%. Acetic acid was the main volatile fatty acid in the fermented straw.     

Enzymatic Saccharification of Pretreated Wheat Straw by T. Reesei Cellulases and A. Niger β-Glucosidase

Three methods of wheat straw treatment (with NaOH, H₂O₂ and butylamine) and its subsequent saccharification by Trichoderma reesei cellulases and Aspergillus niger β-glucosidase are reported. The treatment of straw with NaOH for 1 h in the autoclave (120°C) caused a great loss of the hemicellulose content and a partial removal of lignin, provoking an increase of the cellulose content (from 24% to 69%) in the residue. When the straw was pre-treated with H₂O₂ at 25°C for 20 h, the relative content of cellulose in the straw increased (from 24% to 52%) due to the solubilisation of hemicellulose.

The effect of varying the hydrolysis parameters (enzyme and substrate concentration, temperature and pH) was studied in order to maximise the yield of sugars. Under the best conditions and after 48 h with a mixture of 2:1 (w/w) cellulase/β-glucosidase (with a concentration of 7 and 0.1 U ml^-1, respectively) the native, NaOH-treated and H₂O₂-treated straw material were degraded to reducing sugars for 28%, 89% and 97% respectively.     

Effects of NaOH treatment on condensed tannin contents and gas production kinetics of tree leaves

Effects of NaOH treatment on the crude protein (CP), condensed tannin (CT) and in vitro gas production kinetics of leaves of Arbutus andrachne, Glycyrrhiza glabra L. and wheat straw were determined. Wheat straw, which is tannin-free, was used as the standard. The NaOH treatment was completed by pulverization of samples with 0, 20, 40, 60 and 80 g/L of NaOH solution in the proportion of 1 L of solution to 1 kg of sample. Gas production was determined at 3, 6, 12, 24, 48, 72 and 96 h of incubation. NaOH treatment linearly decreased (P<0.001) the CT contents of leaves of Arbutus andrachne and Glycyrrhiza glabra L. whereas NaOH treatment had no effect on the CP contents of Arbutus andrachne, Glycyrrhiza glabra L. and wheat straw. The 80 g/L NaOH treatment reduced the CT content of leaves of Arbutus andrachne and Glycyrrhiza glabra L. by 59.6% and 86.7%, respectively. NaOH treatment linearly decreased (P<0.01) gas production rate of Arbutus andrachne although it linearly increased (P<0.01) gas production rate of wheat straw. In contrast, NaOH treatment had no effect on gas production rate of leaves of Glycyrrhiza glabra L. NaOH treatment linearly decreased (P<0.001) potential gas production of leaves of Arbutus andrachne and Glycyrrhiza glabra L. whereas NaOH treatment linearly increased (P<0.001) potential gas production of wheat straw. Treatment with NaOH can be used to improve the nutritive value of tannin-free forages such as straw, but not for tannin-containing leaves.     

Bioconversion of wheat straw to ethanol: Chemical modification,enzymatic hydrolysis,and fermentation

Native wheat straw (WS) was pretreated with various concentrations of H₂SO₄ and NaOH followed by secondary treatments with ethylene diamine (EDA) and NH₄OH prior to enzymatic saccharification. Conversion of the cellulosic component to sugar varied with the chemical modification steps. Treatment solely with alkali yield 51–75% conversion, depending on temperature. Acid treatment at elevated tempeatures showed a substantial decrease in the hemicellulose component, whereas EDA-treated WS (acid pretreated) showed a 69–75% decrease in the lignin component. Acid-pretreated EDA-treated straw yielded a 98% conversion rate, followed by 83% for alkali–NH₄OH treated straws. In other experiments, WS was pretreated with varying concentration of H₂SO₄ or NaOh followed by NH₄OH treatment prior to enzymatic hydrolysis. Pretreatment of straw with 2% NaOH for 4 h coupled to enzymatic hydrolysis yield a 76% conversion of the cellulosic component. Acid–base combination pretreatment yielded only 43% conversions. A reactor column was subsequently used to measure modification–saccharification–fermentation for wheat straw conversion on a larger scale. Thirty percent conversions of wheat straw cellulosics to sugar were observed with subsequent fermentation to alcohol. The crude cellulase preparation yielded considerable quantities of xylose in addition to the glucose. Saccharified materials were fermented directly with actively proliferating proliferating yeast cells without concentration of the sugars.     

Stress relaxation of wood partially non-crystallized using aqueous NaOH solutions

Wood samples (Picea jezoensis Carr.) were treated with aqueous NaOH solutions (0-0.20 concentration fraction, 12 conditions), and bending tests were performed to measure stress relaxation. The relationship between mechanical properties and NaOH concentration is discussed. The relaxation modulus and relaxation rate were divided into three concentration ranges. Both decreased slightly for NaOH concentrations less than 0.10, decreased drastically for concentrations between 0.11 and 0.14, and decreased slightly for concentrations greater than 0.15. The change in relaxation behavior upon NaOH treatment was due to an increase in molecular chain mobility in non-crystallized regions along the microfibril longitudinal axis in wood as well as lignin swelling. Furthermore, the molecular chain response in this region required time; thus, the dependence of crystallinity on the relaxation rate was apparent in the long time region.     

Extraction of hemicellulose from ryegrass straw for the production of glucose isomerase and use of the resulting straw residue for animal feed

The hemicellulose fraction of ryegrass straw was extracted with NaOH and used for the production of glucose isomerase by Streptomyces flavogriseus. The level of hemicellulose extracted increased proportionately with increasing NaOH concentration up to about 4%, then the rate of increase slowed down. Hemicellulose extraction was facilitated by the combined application of heat and NaOH. Approximately 15% hemicellulose (12% as pentosan) could be obtained by treating straw with 4% NaOH for either 3 hr at 90°C or 24 hr at room temperature. The highest level (3.04 units/ml culture) of intracellular glucose isomerase was obtained when the organism was grown at 30°C for two days on 2% straw hemicellulose. The organism also produced a high yield of glucose isomerase on xylose or xylan. The NaOH-treated straw residue, after removal of hemicellulose, had approximately 75% higher digestibility and 20% higher feed efficiency for weanling meadow voles than untreated straw. Thus, the residue could be used as animal feed. A process for the production of glucose isomerase and animal feed from ryegrass straw was also proposed.     

Evaluation of chemical pretreatment for enzymatic solubilization of rice straw

Summary Rice straw was treated with NaOH, peracetic acid (PA), and sodium chlorite (NaClO₂). Quantitative changes in the composition of the treated straw, crystallinity of the treated straw and extracted cellulose, and susceptibility of the treated straw to Trichoderma reesei cellulase were studied. The alkali treatment resulted in a remarkable decrease in hemicellulose as well as lignin. Consequently, the recovery of residual straw after NaOH treatment was lowest among the three chemical reagents evaluated. The treatment with PA or NaCIO₂ resulted in a slight loss in hemicellulose and cellulose in the straw. The three chemical treatments caused little or no breakdown of the crystalline structure of cellulose in the straw. The treated straw was solubilized with the culture filtrate of T. reesei. The degree of enzymatic solubilization relative to the amount of residual straw was 69% after treatment with 0.25 N NaOH, 42% after treatment with 20% PA, and 50% after treatments with NaClO₂ (twice). The degree of enzymatic solubilization relative to the amount of the untreated straw, however, was 30% after treatment with 0.25 N NaOH, 32% after treatment with 20% PA, and 37% after treatments with NaClO₂ (twice).     

Mild alkaline/oxidative pretreatment of wheat straw

A new mild alkaline/oxidative pretreatment of wheat straw prior to enzymic hydrolysis was carried out. It consists of a first alkaline (1% NaOH for 24 h) step, which mainly solubilises hemicellullose and renders the material more accessible to further chemical attack, and a second alkaline/oxidative step (1% NaOH and 0·3% H₂O₂ for 24 h), which solubilises and oxidises lignin to minor polluting compounds. The entire process was carried out at low temperature (25–40°C) using a low concentration of chemicals, resulting in a relatively low cost and waste liquors containing only trace amounts of dangerous pollutants derived from lignin. Recovery of cellulose after the double pretreatment reached 90% of that contained in the starting material, with a concomitant 81% degradation of lignin. The action of a commercial cellulase on the cellulose obtained produced a syrup with a high concentration of reducing sugars (220 mg/ml), of which a large percentage was glucose.     

Review article: The alkali treatment of straws

The Beckmann method of alkali treatment consists of soaking straw in dilute alkali solutions for 24 hours and then washing it with clean water. Straw digestibility is increased from about 40 to about 70%. While this process has been known for 50 years, it has not been much used because treatment costs are too high. Also, it cannot be industrialised. The spray process, in which the straw is wetted with an alkali solution (Wilson and Pigden, 1964) is an improvement from both points of view, but poses nutritional problems since straw is not washed after treatment. So great is the potential, however, that within the last seven years nearly 100 research reports have been published on the production and use of spray-treated straw. These reports are reviewed in this article.Straws are poorly digested by ruminants because of their high cell-wall content. Alkali treatment disrupts the cell-wall by dissolving hemicellulose, lignin and silica, by hydrolysing uronic and acetic acid esters and by swelling cellulose. Digestibility in vitro increases from about 40 to about 80% with $\text{10}\text{g NaOH}\text{100}\text{g straw}$. Equally large increases are not, however, obtained in vivo because of unreacted alkali and/or high sodium concentration. Several hypotheses concerning the depression of digestibility in vivo are reviewed. In general $\text{3–6}\text{g NaOH}\text{100}\text{g straw}$ is the optimum. For maximum effectiveness the volume of the alkali solution must be between 50 and $\text{200}\text{ml}\text{100}\text{g straw}$. The usefulness of neutralising unreacted alkali has not yet been determined. Crop and industrial residues with lower initial digestibility than straw (e.g. paddy hulls, bagasse and some types of sawdust) are usually still too poor after treatment (digestibility 30–50%) to be useful feeds. Pressure and temperature increase the effectiveness of alkali, but add to the cost of treatment. The pelleting of treated straws probably increases the effectiveness of alkali treatment. The length of time treated straw is allowed to “cure” before being fed does not affect its digestibility. Chemicals other than NaOH, like chlorine, ammonia and peroxides, are also effective in treating straws but are more expensive and/or more difficult to apply.Animal feeding experiments with sprayed straw have shown its utility for livestock normally fed poor straw diets, in high concentrate diets for growing, finishing and milking stock, and as an extender of silage. The factors affecting the degree of improvement to be expected in digestibility, growth and production from the treatment of straw need to be identified and studied.In spite of its high pH and Na content, sprayed straw has not been found to cause any health problems in livestock when treatment is in the range of 3–8 g NaOH/100 g straw. The extra sodium is excreted in the urine. Water intake and urine volume increase. Milk composition is unaffected.Several factories producing alkali spray-treated straw-based diets in pelleted form are already in operation in Europe. The process is briefly described.     

Alkali-induced conversion of beta-chitin to alpha-chitin

Crystal conversion of beta-chitin to alpha-chitin by aq. NaOH treatment was studied for a highly crystalline beta-chitin sample from diatom spine. The minimum NaOH concentration to cause swelling was between 25% and 30% w/w. The alkali-swollen material was poorly crystalline and was regenerated as alpha-chitin on washing with water. This conversion caused total collapse of the original microfibrillar morphology. These features are similar to those of 7 N-8 N HCl treatment reported earlier, but alkali treatment was free from depolymerization or deacetylation.     

Modeling alkali consumption and digestibility improvement from alkaline treatment of wheat straw

The alkali consumption during alkaline treatment of wheat straw at ambient temperature was measured as a function of time, solids concentration, and alkali concentration. The maximum measured alkali consumption was 5.5 g NaOH/100 g TS over a period of 30 days of treatment. Chemical functional groups (e.g., acyl and carboxyl groups) were measured and compared with the observed alkali consumption. The kinetics of alkali consumption were studied and a model was developed which predicts alkali consumption reasonably well. Use of this model was made to predict biodegradability of alkali-treated wheat straw, since a strong correlation was found to exist between alkali consumption and observed biodegradability. The method of bioconversion used was anaerobic digestion for methane production.     

Pretreatment of cellulosic wastes to increase enzyme reactivity

The enzymatic hydrolysis of cellulose to glucose is generally a slow reaction. Different pretreatments, such as ball milling to a ?200 mesh or swelling in 1–2% NaOH are reported to increase the reactivity considerably. In this work a fiber fraction from cattle manure was treated in an autoclave for 5–30 min at temperatures ranging from 130–200°C. The reactivity of the cellulose, measured by incubating samples with a commercial cellulase preparation for one hour at 50°C and pH 4.8, was increased by a factor of 4–6 compared to NaOH treatment and 10–12 compared to untreated fiber. The increased reaction rate is probably mostly due to an increase in cellulose availability to enzymatic attack, as structural hemicellulose is hydrolyzed and removed during the treatment. Sugars, produced by hemicellulosis, hydrolysis, will react further to give caramelization products. These side reactions were shown to be suppressed by short treatment times. The treated fiber could support growth of a mixed culture of Trichoderma viride and Candida utilis only after washing, indicating the formation of water soluble inhibitory products during treatment. The treatment with high-temperature steam can probably be used also with other cellulosic materials to increase reactivity. This may be an attractive way to prepare low-valued wastes such as manure fibers, straw, stalks, or corn cobs for fermentation processes to increase the protein content or for use directly as ruminant animal feed.     

Processing cereal straws by steam explosion in a pilot plant to enhance digestibility in ruminants

Wheat, barley and oat straws were treated by steam explosion (SE) and then washed with 50g/l NaOH solution. The SE treatment was optimized at batch scale on the basis of carbohydrate recovery. Stocks of fodder (300kg) were produced at 198 degrees C for 2.5min by a continuous reactor and used for in vivo digestibility tests carried out on sheep. The flow-sheet and the mass balances were obtained for the entire process. For the three straws, the water consumption has been 7.3kg/kg of straw. To delignify and improve the digestibility of the straws, 20g of NaOH/kg straw was used. The yield of fodder, lignin and hemicellulose is dependant on the nature of the starting straw. Delignified fodder (insoluble fraction) can be produced with a yield of 0.64, 0.59, 0.55, respectively, from wheat, barley and oat straw. SE improved the digestibility of the straw by 25%; alkaline washing further increased it by 9%. Balanced rations containing, on a DM basis, 1/4 of treated straw, had digestibility coefficients similar to those of commercial rations based on alfalfa.     

Author index volume 65

Spores of Bacillus subtilis 168 were apparently fully inactivated by exposure to 2% (w/v) glutaraldehyde for 20 h but a few spores could be revived by further treatment with 10-100 mM NaOH. A similar effect was found with spores from a range of Bacillus species. A minimum concentration of 5% (w/v) glutaraldehyde was required to prevent the alkali-induced reactivation. The implications of these results for the use of glutaraldehyde as a sporicidal agent are discussed.     

Effect of alkali-treated rice straw supplemented with spent tea leaf and thyroprotein on milk yield,milk composition and certain physiological parameters of dairy cows

Thirty-six Friesian cows in mid-lactation were used to compare the effects of nine treatments.Rice straw untreated (US), or treated with 40 g NaOH in 1.5 l H₂ O kg^?1 straw (TS), was given ad libitum with a concentrate mixture containing 0 or 20% spent tea leaf (STL₀ and STL₂₀) and with or without thyroprotein (TP₁₀ or TP₀). The control treatment (C) represented the standard feeding practice adopted on the farm.The NaOH treatment increased milk yield by 22%, but lowered the butterfat content of milk by 6%. Milk yield and milk composition were not affected by spent tea leaf supplementation. Thyroprotein had no significant effect on milk yield or composition during the experimental period but depressed yield significantly after its withdrawal from the diet. It increased rectal temperature by 0.5° C and heart rate by 5 beats min^?1. All animals on thyroprotein lost body condition.It is concluded that alkali-treated straw is a suitable source of roughage for low yielding cows but its use in practice may be uneconomic. Spent tea leaf is a suitable source of protein for ruminants at the 8% level of substitution in the total diet DM. Thyroprotein feeding is not economical under Sri Lanka conditions and is not recommended for animals on straw diets.