Can Alkaline Hydrolysis of ?-HCH Serve as a Model Reaction to Study Its Aerobic Enzymatic Dehydrochlorination by LinA?
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ABSTRACT:
SUBMITTER: Kannath S
PROVIDER: S-EPMC6929183 | biostudies-literature | 2019 Nov
REPOSITORIES: biostudies-literature
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Project description:Background:Enzymatic hydrolysis is a major step for cellulosic ethanol production. A thorough understanding of enzymatic hydrolysis is necessary to help design optimal conditions and economical systems. The original HCH-1 (Holtzapple-Caram-Humphrey-1) model is a generalized mechanistic model for enzymatic cellulose hydrolysis, but was previously applied only to the initial rates. In this study, the original HCH-1 model was modified to describe integrated enzymatic cellulose hydrolysis. The relationships between parameters in the HCH-1 model and substrate conversion were investigated. Literature models for long-term (>?48 h) enzymatic hydrolysis were summarized and compared to the modified HCH-1 model. Results:A modified HCH-1 model was developed for long-term (>?48 h) enzymatic cellulose hydrolysis. This modified HCH-1 model includes the following additional considerations: (1) relationships between coefficients and substrate conversion, and (2) enzyme stability. Parameter estimation was performed with 10-day experimental data using ?-cellulose as substrate. The developed model satisfactorily describes integrated cellulose hydrolysis data taken with various reaction conditions (initial substrate concentration, initial product concentration, enzyme loading, time). Mechanistic (and semi-mechanistic) literature models for long-term enzymatic hydrolysis were compared with the modified HCH-1 model and evaluated by the corrected version of the Akaike information criterion. Comparison results show that the modified HCH-1 model provides the best fit for enzymatic cellulose hydrolysis. Conclusions:The HCH-1 model was modified to extend its application to integrated enzymatic hydrolysis; it performed well when predicting 10-day cellulose hydrolysis at various experimental conditions. Comparison with the literature models showed that the modified HCH-1 model provided the best fit.
| S-EPMC6378734 | biostudies-literature
Project description:BACKGROUND:A two-stage chemical pretreatment of corn stover is investigated comprising an NaOH pre-extraction followed by an alkaline hydrogen peroxide (AHP) post-treatment. We propose that conventional one-stage AHP pretreatment can be improved using alkaline pre-extraction, which requires significantly less H2O2 and NaOH. To better understand the potential of this approach, this study investigates several components of this process including alkaline pre-extraction, alkaline and alkaline-oxidative post-treatment, fermentation, and the composition of alkali extracts. RESULTS:Mild NaOH pre-extraction of corn stover uses less than 0.1 g NaOH per g corn stover at 80°C. The resulting substrates were highly digestible by cellulolytic enzymes at relatively low enzyme loadings and had a strong susceptibility to drying-induced hydrolysis yield losses. Alkaline pre-extraction was highly selective for lignin removal over xylan removal; xylan removal was relatively minimal (~20%). During alkaline pre-extraction, up to 0.10 g of alkali was consumed per g of corn stover. AHP post-treatment at low oxidant loading (25 mg H2O2 per g pre-extracted biomass) increased glucose hydrolysis yields by 5%, which approached near-theoretical yields. ELISA screening of alkali pre-extraction liquors and the AHP post-treatment liquors demonstrated that xyloglucan and ?-glucans likely remained tightly bound in the biomass whereas the majority of the soluble polymeric xylans were glucurono (arabino) xylans and potentially homoxylans. Pectic polysaccharides were depleted in the AHP post-treatment liquor relative to the alkaline pre-extraction liquor. Because the already-low inhibitor content was further decreased in the alkaline pre-extraction, the hydrolysates generated by this two-stage pretreatment were highly fermentable by Saccharomyces cerevisiae strains that were metabolically engineered and evolved for xylose fermentation. CONCLUSIONS:This work demonstrates that this two-stage pretreatment process is well suited for converting lignocellulose to fermentable sugars and biofuels, such as ethanol. This approach achieved high enzymatic sugars yields from pretreated corn stover using substantially lower oxidant loadings than have been reported previously in the literature. This pretreatment approach allows for many possible process configurations involving novel alkali recovery approaches and novel uses of alkaline pre-extraction liquors. Further work is required to identify the most economical configuration, including process designs using techno-economic analysis and investigating processing strategies that economize water use.
| S-EPMC3997815 | biostudies-literature
Project description:BackgroundDue to the intact structure of lignocellulosic biomass, pretreatment was a prerequisite to improve the enzymatic hydrolysis by disrupting the recalcitrant lignocellulose and increasing the accessibility of cellulose to enzyme. In this study, an alkaline hydrogen peroxide (AHP) pretreatment of sugarcane bagasse with various loadings of H2O2 (1.25-6.25 wt%) at temperatures of 60-160 °C was proposed to degrade hemicellulose/lignin and improve the enzymatic digestibility.ResultsIt was found that increasing H2O2 loadings during pretreatment lead to the enhancement of substrate digestibility, whereas the alkali (only NaOH)-pretreated solid generated higher glucose yield than that pretreated under AHP pretreatment with lower loading of H2O2. This enhancement of enzymatic digestibility was due to the degradation of hemicellulose and lignin. Furthermore, Tween 80 was added to promote enzymatic digestibility, however, the increased yields were different with various substrates and hydrolysis time. The highest glucose yield of 77.6% was obtained after pretreatment at 160 °C for 60 min with 6.25% H2O2 and the addition of Tween 80, representing 89.1% of glucose in pretreated substrate.ConclusionsThis study demonstrated that the AHP pretreatment could greatly enhance the enzymatic saccharification. The addition of Tween 80 played remarkable performances in promoting the glucose yield during enzymatic hydrolysis by stabilizing and protecting the enzyme activity. This study provided an economical feasible and gradual process for the generation of glucose, which will be subsequently converted to bioethanol and bio-chemicals.
| S-EPMC6498686 | biostudies-literature
Project description:BackgroundDuring cellulosic ethanol production, cellulose hydrolysis is achieved by synergistic action of cellulase enzyme complex consisting of multiple enzymes with different mode of actions. Enzymatic hydrolysis of cellulose is one of the bottlenecks in the commercialization of the process due to low hydrolysis rates and high cost of enzymes. A robust hydrolysis model that can predict hydrolysis profile under various scenarios can act as an important forecasting tool to improve the hydrolysis process. However, multiple factors affecting hydrolysis: cellulose structure and complex enzyme-substrate interactions during hydrolysis make it diffucult to develop mathematical kinetic models that can simulate hydrolysis in presence of multiple enzymes with high fidelity. In this study, a comprehensive hydrolysis model based on stochastic molecular modeling approch in which each hydrolysis event is translated into a discrete event is presented. The model captures the structural features of cellulose, enzyme properties (mode of actions, synergism, inhibition), and most importantly dynamic morphological changes in the substrate that directly affect the enzyme-substrate interactions during hydrolysis.ResultsCellulose was modeled as a group of microfibrils consisting of elementary fibrils bundles, where each elementary fibril was represented as a three dimensional matrix of glucose molecules. Hydrolysis of cellulose was simulated based on Monte Carlo simulation technique. Cellulose hydrolysis results predicted by model simulations agree well with the experimental data from literature. Coefficients of determination for model predictions and experimental values were in the range of 0.75 to 0.96 for Avicel hydrolysis by CBH I action. Model was able to simulate the synergistic action of multiple enzymes during hydrolysis. The model simulations captured the important experimental observations: effect of structural properties, enzyme inhibition and enzyme loadings on the hydrolysis and degree of synergism among enzymes.ConclusionsThe model was effective in capturing the dynamic behavior of cellulose hydrolysis during action of individual as well as multiple cellulases. Simulations were in qualitative and quantitative agreement with experimental data. Several experimentally observed phenomena were simulated without the need for any additional assumptions or parameter changes and confirmed the validity of using the stochastic molecular modeling approach to quantitatively and qualitatively describe the cellulose hydrolysis.
| S-EPMC3664589 | biostudies-literature
Project description:Phosphate and sulfate esters have important roles in regulating cellular processes. However, while there has been substantial experimental and computational investigation of the mechanisms and the transition states involved in phosphate ester hydrolysis, there is far less work on sulfate ester hydrolysis. Here, we report a detailed computational study of the alkaline hydrolysis of diaryl sulfate diesters, using different DFT functionals as well as mixed implicit/explicit solvation with varying numbers of explicit water molecules. We consider the impact of the computational model on computed linear free-energy relationships (LFER) and the nature of the transition states (TS) involved. We obtain good qualitative agreement with experimental LFER data when using a pure implicit solvent model and excellent agreement with experimental kinetic isotope effects for all models used. Our calculations suggest that sulfate diester hydrolysis proceeds through loose transition states, with minimal bond formation to the nucleophile and bond cleavage to the leaving group already initiated. Comparison to prior work indicates that these TS are similar in nature to those for the alkaline hydrolysis of neutral arylsulfonate monoesters or charged phosphate diesters and fluorophosphates. Obtaining more detailed insights into the transition states involved assists in understanding the selectivity of enzymes that hydrolyze these reactions.
| S-EPMC7304899 | biostudies-literature
Project description:Complex substrates, like proteins, carbohydrates, and lipids, are major components of domestic wastewater, and yet their degradation in biofilm-based wastewater treatment technologies, such as aerobic granular sludge (AGS), is not well understood. Hydrolysis is considered the rate-limiting step in the bioconversion of complex substrates, and as such, it will impact the utilization of a large wastewater COD (chemical oxygen demand) fraction by the biofilms or granules. To study the hydrolysis of complex substrates within these types of biomass, this paper investigates the anaerobic activity of major hydrolytic enzymes in the different sludge fractions of a full-scale AGS reactor. Chromogenic substrates were used under fully mixed anaerobic conditions to determine lipase, protease, α-glucosidase, and β-glucosidase activities in large granules (>1 mm in diameter), small granules (0.2-1 mm), flocculent sludge (0.045-0.2 mm), and bulk liquid. Furthermore, composition and hydrolytic activity of influent wastewater samples were determined. Our results showed an overcapacity of the sludge to hydrolyze wastewater soluble and colloidal polymeric substrates. The highest specific hydrolytic activity was associated with the flocculent sludge fraction (1.5-7.5 times that of large and smaller granules), in agreement with its large available surface area. However, the biomass in the full-scale reactor consisted of 84% large granules, making the large granules account for 55-68% of the total hydrolytic activity potential in the reactor. These observations shine a new light on the contribution of large granules to the conversion of polymeric COD and suggest that large granules can hydrolyze a significant amount of this influent fraction. The anaerobic removal of polymeric soluble and colloidal substrates could clarify the stable granule formation that is observed in full-scale installations, even when those are fed with complex wastewaters. KEY POINTS: • Large and small granules contain >70% of the hydrolysis potential in an AGS reactor. • Flocculent sludge has high hydrolytic activity but constitutes <10% VS in AGS. • AGS has an overcapacity to hydrolyze complex substrates in domestic wastewater.
| S-EPMC8390406 | biostudies-literature
Project description:Porphyra sensu lato is one of the most economically significant and widely cultured and consumed algae in the world. Porphyra species present excellent nutraceutic properties due to their bioactive compounds (BACs). This research aimed to find the most efficient aqueous extraction method for BACs by examining alkaline and enzymatic hydrolysis. Alkaline hydrolysis with 2.5% sodium carbonate (SC) and at 80 °C proved optimal for extracting all BACs (phycobiliproteins, soluble proteins, polyphenols, and carbohydrates) except mycosporine-like amino acids (MAAs), which were best extracted with water only, and at 80 °C. Enzymatic hydrolysis, particularly with the 'Miura' enzymatic cocktail (cellulase, xylanase, glycoside hydrolase, and β-glucanase), showed superior results in extracting phycoerythrin (PE), phycocyanin (PC), soluble proteins, and carbohydrates, with increases of approximately 195%, 510%, 890%, and 65%, respectively, compared to the best alkaline hydrolysis extraction (2.5% SC and 80 °C). Phenolic content analysis showed no significant difference between the 'Miura' cocktail and 2.5% SC treatments. Antioxidant activity was higher in samples from alkaline hydrolysis, while extraction of MAAs showed no significant difference between water-only and 'Miura' treatments. The study concludes that enzymatic hydrolysis improves the efficiency of BACs extraction in P. linearis, highlighting its potential for the nutraceutical industry, and especially with respect to MAAs for topical and oral UV-photoprotectors.
| S-EPMC11204741 | biostudies-literature
Project description:A maize (Zea mays L. subsp. mays) diversity panel consisting of 26 maize lines exhibiting a wide range of cell-wall properties and responses to hydrolysis by cellulolytic enzymes was employed to investigate the relationship between cell-wall properties, cell-wall responses to mild NaOH pre-treatment, and enzymatic hydrolysis yields. Enzymatic hydrolysis of the cellulose in the untreated maize was found to be positively correlated with the water retention value, which is a measure of cell-wall susceptibility to swelling. It was also positively correlated with the lignin syringyl/guaiacyl ratio and negatively correlated with the initial cell-wall lignin, xylan, acetate, and p-coumaric acid (pCA) content, as well as pCA released from the cell wall by pre-treatment. The hydrolysis yield following pre-treatment exhibited statistically significant negative correlations to the lignin content after pre-treatment and positive correlations to the solubilized ferulic acid and pCA. Several unanticipated results were observed, including a positive correlation between initial lignin and acetate content, lack of correlation between acetate content and initial xylan content, and negative correlation between each of these three variables to the hydrolysis yields for untreated maize. Another surprising result was that pCA release was negatively correlated with hydrolysis yields for untreated maize and, along with ferulic acid release, was positively correlated with the pre-treated maize hydrolysis yields. This indicates that these properties that may negatively contribute to the recalcitrance in untreated cell walls may positively contribute to their deconstruction by alkaline pre-treatment.
| S-EPMC4493778 | biostudies-literature
Project description:BackgroundLignocellulosic biomass has been commonly regarded as a potential feedstock for the production of biofuels and biochemicals. High sugar yields and the complete bioconversion of all the lignocellulosic sugars into valuable products are attractive for the utilization of lignocelluloses. It is essential to pretreat and hydrolyze lignocelluloses at high solids loadings during industrial processes, which is more economical and environmentally friendly as capital cost, energy consumption, and water usage can be reduced. However, oligosaccharides are inevitably released during the high solids loading enzymatic hydrolysis and they are more recalcitrant than monosaccharides for microorganisms.ResultsA fed-batch enzymatic hydrolysis of corn stover pretreated by the sodium hydroxide-methanol solution (SMs) at high solids loading was demonstrated to reach the high concentrations and yields of fermentable sugars. Glucose, xylose, cello-oligosaccharides, and xylo-oligosaccharides achieved 146.7 g/L, 58.7 g/L, 15.6 g/L, and 24.7 g/L, respectively, when the fed-batch hydrolysis was started at 12% (w/v) solids loading, and 7% fresh substrate and a standardized blend of cellulase, β-glucosidase, and hemicellulase were fed consecutively at 3, 6, 24, and 48 h to achieve a final solids loading of 40% (w/v). The total conversion of glucan and xylan reached 89.5% and 88.5%, respectively, when the oligosaccharides were taken into account. Then, a fed-batch culture on the hydrolysates was investigated for lipid production by Cutaneotrichosporon oleaginosum. Biomass, lipid content, and lipid yield were 50.7 g/L, 61.7%, and 0.18 g/g, respectively. The overall consumptions of cello-oligosaccharides and xylo-oligosaccharides reached 74.1% and 68.2%, respectively.ConclusionsHigh sugars concentrations and yields were achieved when the enzyme blend was supplemented simultaneously with the substrate at each time point of feeding during the fed-batch enzymatic hydrolysis. Oligosaccharides were co-utilized with monosaccharides during the fed-batch culture of C. oleaginosum. These results provide a promising strategy to hydrolyze alkaline organosolv-pretreated corn stover into fermentable sugars with high concentrations and yields for microbial lipid production.
| S-EPMC6977323 | biostudies-literature
Project description:The semiempirical PM3 method, calibrated against ab initio HF/6-31+G(d) theory, has been used to elucidate the reaction of 1, 2-dichloroethane (DCE) with the carboxylate of Asp-124 at the active site of haloalkane dehalogenase of Xanthobacter autothropicus. Asp-124 and 13 other amino acid side chains that make up the active site cavity (Glu-56, Trp-125, Phe-128, Phe-172, Trp-175, Leu-179, Val-219, Phe-222, Pro-223, Val-226, Leu-262, Leu-263, and His-289) were included in the calculations. The three most significant observations of the present study are that: (i) the DCE substrate and Asp-124 carboxylate, in the reactive ES complex, are present as an ion-molecule complex with a structure similar to that seen in the gas-phase reaction of AcO- with DCE; (ii) the structures of the transition states in the gas-phase and enzymatic reaction are much the same where the structure formed at the active site is somewhat exploded; and (iii) the enthalpies in going from ground states to transition states in the enzymatic and gas-phase reactions differ by only a couple kcal/mol. The dehalogenase derives its catalytic power from: (i) bringing the electrophile and nucleophile together in a low-dielectric environment in an orientation that allows the reaction to occur without much structural reorganization; (ii) desolvation; and (iii) stabilizing the leaving chloride anion by Trp-125 and Trp-175 through hydrogen bonding.
| S-EPMC22940 | biostudies-literature