Loss Of Xylose Research Articles

Genetically Engineering Escherichia coli to Produce Xylitol from Corncob Hydrolysate without Lime Detoxification.

Before fermentation with hemicellulosic hydrolysate as a substrate, it is generally necessary to detoxify the toxic substances that are harmful to microorganism growth. Cyclic AMP receptor protein (CRP) is a global regulator, and mutation of its key sites may have an important impact on E. coli virulence tolerance. Using corncob hydrolysate without ion-exchange or lime detoxification as the substrate, shake flask fermentation experiments showed that CRP mutant IS5-dG (I112L, T127G, A144T) produced 18.4 g/L of xylitol within 34 h, and the OD600 was 9.7 at 24 h; these values were 41.5% and 21.3% higher than those of the starting strain, IS5-d, respectively. This mutant produced 82 g/L of xylitol from corncob hydrolysate without ion-exchange or lime detoxification during fed-batch fermentation in a 15-L bioreactor, with a productivity of 1.04 g/L/h; these values were 173% and 174% higher than the starting strain, respectively. To our knowledge, this is the highest xylitol concentration and productivity produced by microbial fermentation using completely non-detoxified hemicellulosic hydrolysate as the substrate to date. This study also showed that alkali neutralization, high temperature sterilization, and fermentation of the hydrolysate had important effects on the xylose loss rate and xylitol production.

A two-stage heating process with low energy consumption for treatment of preliminary desalinated impregnation effluent to produce xylose

Production of xylose from preliminary desalinated impregnation effluent (PDSIE) containing hemicellulose is an economical way of the effluent treatment. However, due to the high alkali and salt contents in PDSIE, its resource utilization is limited in the developed treatment process for pulp-impregnated effluent that has been deeply desalted. In this study, based on analysis of reaction rate and kinetic characteristics of pre-hydrolysis, a two-stage pre-hydrolysis heating process including heating at a high temperature for a short time followed by slow cooling to enzymatic hydrolysis temperature (HCP) with low energy consumption was developed for PDSIE. The heating time was shortened from 2 h to 0.5 h, the inhibition of conversion from hemicellulose to xylose by the high alkali and salt contents in PDSIE was alleviated, and the loss of xylose was reduced by HCP, compared to those under the constant temperature heating conditions. The heating cost based on coal consumption was reduced by 75.54% in HCP according to the preliminary economic analysis. Based on xylose yield and energy-saving results, the temperature in the first stage of HCP was determined to be 120 °C, and the cooling time in the second stage was determined to be 6 h.

Improvement of bioethanol production using a new fermentation system: The process analysis and micro-mechanisms study

Rice straw based bioethanol is a clean alternative energy source to alleviate the energy crisis and greenhouse gases emission. However, the efficient alkali pretreatment of rice straw for bioethanol production will generate several fermentation inhibitors, such as ferulic acid, which could inhibit bioethanol fermentation. Therefore, an adsorbent called Air Environment-Prepared Adsorbent at 250 ℃ (AEPA250) was prepared using the enzymatically hydrolyzed residue of rice straw to detoxify ferulic acid in this study for enhancing the subsequent bioethanol production. Analysis of the mass balance showed that ferulic acid detoxification by AEPA250 had a high removal efficiency of 94.393% with a low glucose and xylose loss of 2.532% and 8.219%, respectively. A higher 277.551 mg/L bioethanol concentration and 76.005% glucose consumption rate in the SSBP system were obtained compared to non-detoxified sample. Furthermore, proteomics analysis indicated that certain metabolic pathways of TCA cycle and ribosome pathway as well as various coded proteins of ACO1, MRP2, RPL24B, MRPL33, RPL32, RPL39, RPS17B, RPS19A, RPS26A and ATP5 contributed to ferulic acid detoxification in the SSBP system. The findings of this study may help develop efficient pretreatment methods, detoxification strategies and engineering yeast strains for improving bioethanol production in the future.

Lysine Production by Dry Biorefining of Wheat Straw and Cofermentation of Corynebacterium glutamicum.

A preliminary study shows that lysine production from lignocellulose feedstock is feasible, but the conversion of xylose in lignocellulose to lysine remains unsolved. Two technical barriers are responsible for the remaining xylose conversion: one is the xylose loss into the wastewater stream of the biorefinery processing chain, and the other is the lack of efficient lysine-producing strain with xylose utilization. Here, we conducted a new biorefinery approach of consequent dry acid pretreatment and biodetoxification, resulting in zero wastewater generation and then well-preserved xylose. To provide the lysine-producing strain with xylose utilization, we modified the Corynebacterium glutamicum by establishing the xylose assimilation pathway and improving the NADPH cofactor regeneration. The combinational modification of biorefinery processing and strain development led to 31.3 g/L of lysine production with a yield of 0.23 g lysine per gram of wheat straw derived sugars. This study provides a practical method for upgraded lysine production from lignocellulose for future industrial applications.

Selective single-stage xylan-to-xylose hydrolysis and its effect on enzymatic digestibility of energy crops giant reed and cardoon for bioethanol production

The low temperature dilute sulfuric acid hydrolysis of two potential European energy crops − giant reed and cardoon have been studied for separation and valorization of total hemicellulosic carbohydrates prior to fractionation of cellulose and lignin, viewing as entry point to integrated biorefining scheme. The combined severity factor (CSF) was used to assess, on comparative basis, the selectivity and efficiency of xylan-to-xylose conversion and its effect on enzymatic digestibility of residual cellulose. The reliable quantitative correlations have been established for the first time between the CSF and the main outputs of acid and enzymatic hydrolysis. Xylose recovery of ca. 94% and 86% (as max.) was achieved at CSF 1.90 and 1.97, respectively for giant reed and cardoon, with formation of ca. 2.4% glucose, 1.1% furfural, 0.5% HMF and 3.2% acetic acid. Under these CSF levels, the enzymatic digestibility of residual cellulose was improved only up to 0.75 and 0.80 (75% and 80% total Glc recovery, respectively for giant reed and cardoon), vs. 0.09 and 0.19 for untreated biomass. The digestibility was further improved to 0.93 and 0.98 with increase in hydrolysis severity up to CSF 2.44, while substantial loss of xylose (ca. 40%) and furans formation (ca. 5% furfural) was found. The essential effect of cardoon stalk morphology on uniformity and modeling of biorefining separation processes was noted.

Separation of xylose and glucose using an integrated membrane system for enzymatic cofactor regeneration and downstream purification

Mixtures of xylose, glucose and pyruvate were fed to a membrane bioreactor equipped with a charged NF membrane (NTR 7450). Value-added products were obtained in the reactor via enzymatic cofactor-dependent catalysis of glucose to gluconic acid and pyruvate to lactic acid, respectively. The initial cofactor (NADH) concentration could be decreased to 10% of the stoichiometric value (relative to glucose) without compromising process time and substrate conversion via i) efficient cofactor regeneration and ii) high retention of cofactor (R=0.98) in the membrane bioreactor. Furthermore, accumulation of xylose (R<0.1), lactic acid (R=0.38) and gluconic acid (R=0.63) was minimized. After separation of the cofactor in the membrane bioreactor, xylose, lactic acid and gluconic acid were separated based on charge repulsion and size exclusion in a sequence of two NF steps. Broad substrate specificity of glucose dehydrogenase (EC 1.1.1.47) (GDH) lead to partial conversion of xylose to xylonic acid, causing some loss of xylose, but the results obtained nevertheless showed that it is possible to build a robust system for conducting enzyme reactions by sequentially regenerating the cofactor and at the same time obtaining valuable products of high purity.

Removal of furan and phenolic compounds from simulated biomass hydrolysates by batch adsorption and continuous fixed-bed column adsorption methods

It has been proposed to remove all potential inhibitors and sulfuric acid in biomass hydrolysates generated from dilute-acid pretreatment of biomass, based on three steps of sugar purification process. This study focused on its first step in which furan and phenolic compounds were selectively removed from the simulated hydrolysates using activated charcoal. Batch adsorption experiments demonstrated that the affinity of activated charcoal for each component was highest in the order of vanillic acid, 4-hydroxybenzoic acid, furfural, acetic acid, sulfuric acid, and xylose. The affinity of activated charcoal for furan and phenolic compounds proved to be significantly higher than that of the other three components. Four separation strategies were conducted with a combination of batch adsorption and continuous fixed-bed column adsorption methods. It was observed that xylose loss was negligible with near complete removal of furan and phenolic compounds, when at least one fixed-bed column adsorption was implemented in the strategy.

Development and characterization of a high-solids deacetylation process

Dilute-acid pretreatment has proven to be a robust means of converting herbaceous feedstock to fermentable sugars. However, it also releases acetic acid, a known fermentation inhibitor, from acetyl groups present in the biomass. A mild, dilute alkaline extraction stage was implemented prior to acid pretreatment to separate acetic acid from the hydrolysate sugar stream. This step, termed deacetylation, improved the glucose and xylose yields from enzymatic hydrolysis and ethanol yields from fermentation of the sugars relative to the control experiments using dilute-acid pretreatment of native corn stover without deacetylation. While promising, deacetylation as it was historically practiced is conducted at low solids loadings, and at fixed conditions. Thus, many questions have been left unanswered, including the relationship between sodium hydroxide and solids loading, and acetate and xylan solubilization, as well as the impact of temperature and residence time on the process efficacy. A central composite experiment was designed to evaluate the impact of solids loading, sodium hydroxide loading, reaction time and temperature during deacetylation on the acetate and xylan solubilization of corn stover. Using the ANOVA test, it became apparent that neither of the responses was significantly impacted by the solids loading, while the reaction time was a minor factor—the responses were largely driven by reaction temperature and the sodium hydroxide loading. Based on the results, we successfully demonstrated the ability to transition the low-solids (10 % w/w) deacetylation process to a higher-solids process (30 % w/w) with minimal impact on the ability to extract acetate from biomass. Conditions were selected to minimize xylose loss during deacetylation, while also removing 70 % of acetyl groups. The impact of selected conditions on the enzymatic hydrolysis and fermentation was further investigated. Evaluation of the whole-process impact demonstrated that despite the upfront reduction in carbohydrate loss during deacetylation, the overall process sugar yields were depressed by the high-solids, low alkali process relative to the historical control. Consequently, ethanol titers were reduced, though strong fermentation performance was still observed, indicating that 70 % acetate removal is sufficient to depress acetic acid concentrations to a level that does not substantially inhibit ethanol fermentation by rZymomo nas.

Removal of acetic acid from simulated hemicellulosic hydrolysates by emulsion liquid membrane with organophosphorus extractants

Selective removal of acetic acid from simulated hemicellulosic hydrolysates containing xylose and sulfuric acid was attempted in a batch emulsion liquid membrane (ELM) system with organophosphorus extractants. Various experimental variables were used to develop a more energy-efficient ELM process. Total operation time of an ELM run with a very small quantity of trioctylphosphine oxide as the extractant was reduced to about a third of those required to attain almost the same extraction efficiency as obtained in previous ELM works without any extractant. Under specific conditions, acetic acid was selectively separated with a high degree of extraction and insignificant loss of xylose, and its purity and enrichment ratio in the stripping phase were higher than 92% and 6, respectively. Also, reused organic membrane solutions exhibited the extraction efficiency as high as fresh organic solutions did. These results showed that the current ELM process would be quite practical.

Effect of steam explosion on waste copier paper alone and in a mixed lignocellulosic substrate on saccharification and fermentation

This study evaluated steam (SE) explosion on the saccharification and simultaneous saccharification and fermentation (SSF) of waste copier paper. SE resulted in a colouration, a reduction in fibre thickness and increased water absorption. Changes in chemical composition were evident at severities greater than 4.24 resulting in a loss of xylose and the production of breakdown products known to inhibit fermentation (particularly formic acid and acetic acid). SE did not improve final yields of glucose or ethanol, and at severities 4.53 and 4.83 reduced yields probably due to the effect of breakdown products and fermentation inhibitors. However, at moderate severities of 3.6 and 3.9 there was an increase in initial rates of hydrolysis which may provide a basis for reducing processing times. Co-steam explosion of waste copier paper and wheat straw attenuated the production of breakdown products, and may also provide a basis for improving SSF of lignocellulose.

W/O 에멀젼의 주요 구성 성분들이 모사 헤미셀룰로오스 가수분해액에 있는 초산의 제거에 미치는 영향

초산은 묽은 산에 의해 처리되어 얻어진 헤미셀룰로오스 가수분해액 중 가장 양이 많을 뿐만 아니라 중대한 에탄올 발효 저해물질이다. 그 가수분해액으로부터 초산을 선택적으로 제거하는데 최적인 분리시스템을 찾기 위하여 모사 헤미셀룰로오스 가수분해액으로 자일로스, 초산과 황산의 혼합물이 선택되었다. 이러한 목적을 이루기 위해 에멀젼형 액막법이 모사 헤미셀룰로오스 가수분해액으로부터 초산을 제거하는데 적용되었다. W/O 에멀젼의 주요 구성 성분들, 즉 아민계 추출제 종류, 계면활성제 조성, 첨가제 종류와 회수제의 종류 및 농도가 모사 헤미셀룰로오스 가수분해액에 있는 초산, 자일로스와 황산의 추출에 미치는 영향이 조사되었다. 특별한 실험 조건에서, 초산의 추출율은 95% 이상이었고, 자일로스의 손실은 미미하였는데, 이것은 현 에멀젼형 액막법이 경제성이 높은 공정이 될 수 있다는 것을 의미한다. Acetic acid is the most abundant and serious ethanol fermentation inhibitor in dilute-acid hemicellulosic hydrolysates. A mixture of xylose, acetic acid and sulfuric acid was chosen as a simulated hemicellulosic hydrolysate so as to find an optimal separation system to selectively remove acetic acid from the hydrolysates. In order to attain the purpose, emulsion liquid membrane was applied to removal of acetic acid from the simulated hemicellulosic hydrolysate. The effects of main constituents of water-in-oil (W/O) emulsion, such as amine extractant type, surfactant composition, additive type, and type and concentration of stripping agent, on extraction of acetic acid, xylose, and sulfuric acid in the simulated hemicellulosic hydrolysate were investigated. Under specific experimental conditions, degree of extraction of acetic acid was higher than 95% while loss of xylose was insignificant, which means that the current emulsion liquid membrane can be an economically feasible process.

Purification of xylose in simulated hemicellulosic hydrolysates using a two-step emulsion liquid membrane process

Purification of xylose in simulated hemicellulosic hydrolysates was attempted using a two-step emulsion liquid membrane (ELM) process. The effects of various experimental variables on extraction of each component in the hydrolysates were investigated in the ELM steps. In the first ELM step, acetic acid could be selectively removed from the hydrolysates and highly enriched in the stripping phase, and loss of xylose was insignificant. In the second ELM step, sulfuric acid could be selectively removed from simulated acetic acid-free hemicellulosic hydrolysates and somewhat enriched in the stripping phase. There was just small loss of xylose, and the final pH of the feed phase approached a pH level suitable for ethanol fermentation. Also, concentration of xylose in the feed phase was attained as an incidental outcome during each ELM run. Conclusively, the two-step ELM process was found to be a promising futuristic technology for purification of sugars in real hemicellulosic hydrolysates.

Pilot-scale ethanol production from rice straw hydrolysates using xylose-fermenting Pichia stipitis

Ethanol was produced at pilot scale from rice straw hydrolysates using a Pichia stipitis strain previously adapted to NaOH-neutralized hydrolysates. The highest ethanol yield was 0.44±0.02gp/gs at an aeration rate of 0.05vvm using overliming-detoxified hydrolysates. The yield with hydrolysates conditioned by ammonia and NaOH was 0.39±0.01 and 0.34±0.01gp/gs, respectively, were achieved at the same aeration rate. The actual ethanol yield from hydrolysate fermentation with ammonia neutralization was similar to that with overliming hydrolysate after taking into account the xylose loss resulting from these conditioning processes. Moreover, the ethanol yield from ammonia-neutralized hydrolysates could be further enhanced by increasing the initial cell density by two-fold or reducing the combined concentration of furfural and 5-hydroxymethyl furfural to 0.6g/L by reducing the severity of operational conditions in pretreatment. This study demonstrated the potential for commercial ethanol production from rice straw via xylose fermentation.

EFEITO DA DESTOXIFICAÇÃO DO HIDROLISADO DE BAGAÇO DE CANA SOBRE A REMOÇÃO DE FENÓIS, A PERDA DE AÇÚCARES E A BIOCONVERSÃO DE XILOSE EM XILITOL

RESUMO: O hidrolisado resultante do processo de hidrolise acida, metodologia comumente empregada para diferentes materiais lignocelulosicos, contem, alem dos acucares, compostos toxicos aos microrganismos. Dentre estes, os compostos fenolicos, tem sido mencionados como inibidores da bioconversao de xilose em xilitol, quer pela sua acao individual ou pela interacao sinergistica entre ele e outros compostos presentes nos hidrolisados como o furfural, hidroximetilfurfural e o acido acetico. Os compostos fenolicos conferem, ainda, ao hidrolisado uma coloracao escura, a qual interfere no processo de “downstrean” do xilitol. Este trabalho avaliou dois procedimentos de destoxificacao do hidrolisado de bagaco de cana-de-acucar: ajuste de pH combinado a adsorcao em carvao ativo e floculacao por polimero de origem vegetal. Os fenois totais e as concentracoes de xilose e glicose foram determinados pelo metodo de Folin-Ciocalteu e atraves de cromatografia liquida, respectivamente. Ambos os tratamentos resultaram em reducao da concentracao de fenois totais. Porem, o uso de carvao ativo resultou em maior perda deste composto (88.5% remocao), o que representa 17.8% de aumento em comparacao ao emprego do polimero. A perda de xilose foi minima (9%) para ambos os tratamentos empregados. O tratamento de ajuste de pH combinado a adsorcao em carvao ativo foi mais eficiente do que o procedimento de floculacao por polimero. Considerando estes ensaios preliminares e tambem o baixo custo e biodegradabilidade do polimero, pesquisas devem ser realizadas para que se estabelecam condicoes mais eficientes do uso deste composto, melhorando a fermentabilidade do hidrolisado e diminuindo a perda de xilose. Palavras-chave: Hidrolisado de bagaco de cana-de-acucar. Destoxificacao. Xilitol. Xilose. Compostos fenolicos. SUMMARY: The hydrolysate resulting from the acid hydrolysis process, a method usually employed for different lignocellulosic materials, contains, besides sugars, compounds that are toxic to the microorganisms. Among them, there are the phenolic compounds, that have inhibitory effect on the xylose-to-xylitol bioconversion, due to their individual action and also to their synergistic interaction with other compounds present in the hydrolysates, such as furfural, hydroxymethylfurfural and acetic acid. Furthermore, the phenolic compounds are responsible for the dark color of the hydrolysate, which interferes in the xilitol downstream process. This work evaluated two procedures of sugarcane bagasse hydrolysate detoxification: pH adjusting with active charcoal adsorption, and a vegetable-origin polymer flocculation technique. The total phenolics and the xylose and glycose concentrations were determined by Folin-Ciocalteu method and liquid chromatography, respectively. Both treatments resulted in reduction of total phenolics concentration. However, the use of active charcoal resulted in higher loss of this compound (88.5% removal), that represents 17.8% increase when comparised to the use of polymer. The xylose loss was minimum (8%) for both used treatments. The treatment using the combination of pH adjustment and active charcoal adsorption was more efficient than the procedure that used vegetable-origin polymer. Considering these preliminary assays and also the polymer low cost and its biodegradability, researches must be performed in order to establish conditions for the most effective use of this compound, improving the hydrolysate fermentability � Doutoranda/CAPES, DEBIQ, EEL-USP. Estrada Municipal do Campinho s/n, CEP �2602-8�0, Lorena

Effect of an Enzyme Additive on Ensilage Losses of Plant Cell Wall in Italian Ryegrass

The effects of an additive containing cell wall degrading enzyme on the ensilage losses were estimated under a farm condition. At ensiling Italian ryegrass with and without the additive, the nylon-net bags containing grasses were placed into each of the silage bulks. The bags were retrieved when the silages were unloaded, and their contents were subjected to the chemical analyses, in vitro digestion with rumen microbes and microscopic observation. Enzyme treatment improved the silage fermentation. The ensilage loss of cell wall components was highest in hemicellulose. In hemicellulosic monosaccharides, the loss of xylose was lowest, followed by uronic acids, arabinose, galactose and glucose. The losses in treated silage were significantly higher than the control silage in almost all the cell wall components. The losses of digestible-dry matter (DM) and digestible-neutral detergent fiber (NDF) in vitro were also significantly higher in the treated silage. The selective degradations of the readily digestible plant tissues (e. g. mesophyll and pith parenchyma) were observed in the treated silage.