Simultaneous Hydrolysis Research Articles

Biohydrogen from cellulosic feedstock: Dilution-to-stimulation approach

Biohydrogen produced from cellulosic feedstock is a promising candidate as a clean energy carrier to meet future energy needs. This work identified a functional consortium that can produce biohydrogen from three cellulosic feedstocks, the Whatman No. 2 filter, xylan, and Chlorella cells, using a novel dilution-to-stimulation approach. By serially diluting an anaerobic inoculum, biohydrogen production increased, peaked at a specific dilution ratio, and then declined. With filter paper tests at 10−9 dilution, the functional consortium was mainly composed of Clostridium sp. clone UC61. At dilutions greater than 10−9, the presence of other heterotrophic strains diminished the ability of consortium to produce hydrogen. In the xylan tests, Clostridium sp. clone UC61, Clostridium sp. BN2, and Bacteroides species comprised the functional consortium. Klebsiella sp. competed with Clostridium sp. for carbon and decreased hydrogen production. In the Chlorella cells tests, Bacillus, Acinetobacter, Elusimicrobium, and Clostridium comprised the functional consortium. The loss of Bacillus and Acinetobacter species under high dilution deteriorated the synergistic relationships among different strains. The proposed dilution-to-stimulation approach is effective for isolating functional strains for simultaneous hydrolysis and hydrogen production by cellulosic materials.

Amylolysis of large and small granules of native triticale, wheat and corn starches using a mixture of α-amylase and glucoamylase

Starch granules from different sources differ in their morphology and structure, resulting in diverse amylolysis, which is closely related to bioconversion efficiency of starch to fermentable sugars during ethanol production. In this study, large and small granules were fractionated from native triticale, wheat and corn starches using a centrifugal sedimentation protocol and were hydrolyzed by a granular starch hydrolyzing enzyme at sub-gelatinization temperatures. Native triticale starches had bimodal granule size distribution with highest sizes of 14.9–15.9, 19.1–20.2, and 7.2 μm in diameter for overall, large and small granules, respectively, when compared to those of wheat and corn starches. The large granules in triticale starches contributed higher proportion of the total granules either by number (36–45%) or by weight (93–95%) than did those of wheat and corn starches. Surface pores and internal channels viewed by SEM and CLSM were more visible in large granules than in small granules of triticale, wheat, and corn starches, but corn starch generally displayed more pores and channels than triticale and wheat starches. Large granules contained significantly higher apparent amylose content and higher relative crystallinity than did small granules (23.0–28.5% vs. 12.4–21.0% and 24.9–26.8% vs. 20.8–24.4%, respectively) in all starches. Small granules were hydrolyzed significantly faster than did large granules initially (DH 9.3–18.0%, 24.7–40.3%, 17.7% higher in small granules than in large counterparts of triticale, wheat, and corn starches, respectively, at 55 °C for 1 h), however slowed down in later stage of hydrolysis. The final DH of large and small granules were similar in triticale (82.3–84.5%) and corn (80.8–83.4%) starches except wheat starches (90.8–95.1% in large and 79.6–86.2% in small granules) at 72 h hydrolysis. SEM and CLSM further revealed that the hydrolysis pattern differed between large and small starch granules and among starches, indicating diverse structure of large and small granules existed at macro, micro, and nano levels within and between species and cultivars. The study may be beneficial for precise control of liquefaction and saccharification in simultaneous hydrolysis and fermentation process of ethanol production.

Effect of Initial Sugar Concentration on the Production of L (+) Lactic Acid by Simultaneous Enzymatic Hydrolysis and Fermentation of an Agro-Industrial Waste Product of Pineapple (Ananas comosus) Using Lactobacillus casei Subspecies rhamnosus

Production of lactic acid by fermentation process has been studied from glucose solutions and other sources because of many important reasons: biotechnological production is cheaper than chemical synthesis; production of biodegradable materials from L (+) lactic acid and, the use of nutrient-rich agro-industrial wastes as raw material, which helps to reduce the environmental impact. The goal of this study was to evaluate the effect of sugar concentration of a pineapple liquid waste as the carbon source on the capacity of Lactobacillus casei subspecies rhamnosus to produce lactic acid by simultaneous enzymatic hydrolysis and fermentation. Three different pineapple waste concentrations were evaluated (60, 80 and 100% v/v) from a pineapple juice with 11.3% (m/v) of sugars (sucrose, fructose and glucose). L. casei was able to consume all sugars present within the levels tested, and converted all into lactic acid, showing efficient yields of 0.91 g lactic acid/g sugars. Final lactic acid concentration increased significantly (p<0.05) with the increase of pineapple waste percentage. Maximum lactic acid concentration (102g/L) was achieved with 100% pineapple waste medium. The highest total productivity (4.0g/h) and maximum productivity (4.48 g/L*h) were obtained with 60% pineapple waste medium and it decreased significantly (p<0.05) when 100% was used. Fermentation time increased with the increment of sugars, but it increased considerably with the medium composed of 100% of pineapple waste in comparison with the other two mediums. Pineapple waste represents a good alternative as a cheap carbon source for bacteria growth and production of L (+) lactic acid.

Efficient microwave-assisted production of furfural from C5 sugars in aqueous media catalysed by Brönsted acidic ionic liquids

Small amounts of SO3H-functionalised room temperature synthesized ionic liquids efficiently dehydrate aqueous xylose to furfural under microwave heating at mild reaction conditions. The RT-ionic liquid catalysts were also found to be effective catalysts for the two step one-pot simultaneous hydrolysis and dehydration of a lignocellulosic waste biorefinery-derived syrup enriched in C5 sugar oligomers.

A method for the production of D-tagatose using a recombinant Pichia pastoris strain secreting β-D-galactosidase from Arthrobacter chlorophenolicus and a recombinant L-arabinose isomerase from Arthrobacter sp. 22c

BackgroundD-Tagatose is a natural monosaccharide which can be used as a low-calorie sugar substitute in food, beverages and pharmaceutical products. It is also currently being tested as an anti-diabetic and obesity control drug. D-Tagatose is a rare sugar, but it can be manufactured by the chemical or enzymatic isomerization of D-galactose obtained by a β-D-galactosidase-catalyzed hydrolysis of milk sugar lactose and the separation of D-glucose and D-galactose. L-Arabinose isomerases catalyze in vitro the conversion of D-galactose to D-tagatose and are the most promising enzymes for the large-scale production of D-tagatose.ResultsIn this study, the araA gene from psychrotolerant Antarctic bacterium Arthrobacter sp. 22c was isolated, cloned and expressed in Escherichia coli. The active form of recombinant Arthrobacter sp. 22c L-arabinose isomerase consists of six subunits with a combined molecular weight of approximately 335 kDa. The maximum activity of this enzyme towards D-galactose was determined as occurring at 52°C; however, it exhibited over 60% of maximum activity at 30°C. The recombinant Arthrobacter sp. 22c L-arabinose isomerase was optimally active at a broad pH range of 5 to 9. This enzyme is not dependent on divalent metal ions, since it was only marginally activated by Mg2+, Mn2+ or Ca2+ and slightly inhibited by Co2+ or Ni2+. The bioconversion yield of D-galactose to D-tagatose by the purified L-arabinose isomerase reached 30% after 36 h at 50°C. In this study, a recombinant Pichia pastoris yeast strain secreting β-D-galactosidase Arthrobacter chlorophenolicus was also constructed. During cultivation of this strain in a whey permeate, lactose was hydrolyzed and D-glucose was metabolized, whereas D-galactose was accumulated in the medium. Moreover, cultivation of the P. pastoris strain secreting β-D-galactosidase in a whey permeate supplemented with Arthrobacter sp. 22c L-arabinose isomerase resulted in a 90% yield of lactose hydrolysis, the complete utilization of D-glucose and a 30% conversion of D-galactose to D-tagatose.ConclusionsThe method developed for the simultaneous hydrolysis of lactose, utilization of D-glucose and isomerization of D-galactose using a P. pastoris strain secreting β-D-galactosidase and recombinant L-arabinose isomerase seems to offer an interesting alternative for the production of D-tagatose from lactose-containing feedstock.

Direct conversion of cellulose into sorbitol using dual-functionalized catalysts in neutral aqueous solution

Abstract Direct and selective conversion of cellulose into sorbitol was carried out over dual-functionalized catalysts containing both sulfonate groups and Ru nanoparticles. A high sorbitol yield of 71.1% was obtained in a neutral aqueous solution without an aid of liquid phase acid at an intermediate reaction temperature of 165 °C via synergy of sulfonate groups and Ru sites on the catalyst surface. No deactivation was observed even after 5th repeated reactions. The effect of reaction time, temperature, and the Ru amount was also evaluated for the sorbitol production. The cellulose was decomposed by simultaneous hydrolysis and hydrogenation producing cello-oligomers with partially hydrogenated end groups.

Utilization of Cassava Fibrous Residue for the Production of Glucose and High Fructose Syrup

Abstract Cassava (Manihot esculenta Crantz) is one of the largest sources of starch and sago. The cassava fibrous residue (CFR) contains 61%-63% starch and other polysaccharides and causes major disposal problems. Due to its high starch content, it can be utilized to obtain value-added products. Work was therefore undertaken to obtain glucose syrup by hydrolyzing starch and cellulose with various enzyme treatments, followed by conversion to high fructose syrup. Standardization of various conditions for hydrolysis was determined. A combination of α-amylase and glucoamylase (T1 & T2) resulted in 52.88–54.24% conversion of CFR to glucose. The yield could be enhanced to 58.70–60.00% by adding cellulase enzyme complex, Accellerase 1000, in combination with α-amylase and glucoamylase. Cellulase activity helped to reduce viscosity of the CFR slurry and also improved the starch hydrolysis by α-amylase and glucoamylase. The most suitable enzyme treatment was found to be simultaneous hydrolysis of CFR with glucoamy...

The Atypical Two-component Sensor Kinase Lpl0330 from Legionella pneumophila Controls the Bifunctional Diguanylate Cyclase-Phosphodiesterase Lpl0329 to Modulate Bis-(3′-5′)-cyclic Dimeric GMP Synthesis

A significant part of bacterial two-component system response regulators contains effector domains predicted to be involved in metabolism of bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), a second messenger that plays a key role in many physiological processes. The intracellular level of c-di-GMP is controlled by diguanylate cyclase and phosphodiesterases activities associated with GGDEF and EAL domains, respectively. The Legionella pneumophila Lens genome displays 22 GGDEF/EAL domain-encoding genes. One of them, lpl0329, encodes a protein containing a two-component system receiver domain and both GGDEF and EAL domains. Here, we demonstrated that the GGDEF and EAL domains of Lpl0329 are both functional and lead to simultaneous synthesis and hydrolysis of c-di-GMP. Moreover, these two opposite activities are finely regulated by Lpl0329 phosphorylation due to the atypical histidine kinase Lpl0330. Indeed, Lpl0330 was found to autophosphorylate on a histidine residue in an atypical H box, which is conserved in various bacteria species and thus defines a new histidine kinase subfamily. Lpl0330 also catalyzes the phosphotransferase to Lpl0329, which results in a diguanylate cyclase activity decrease whereas phosphodiesterase activity remains efficient. Altogether, these data present (i) a new histidine kinase subfamily based on the conservation of an original H box that we named HGN H box, and (ii) the first example of a bifunctional enzyme that modulates synthesis and turnover of c-di-GMP in response to phosphorylation of its receiver domain.

Hydrolysis and oxidation on supported phosphate catalyst for decomposition of SF6

In order to decompose SF6, AlPO4/γ-alumina catalysts were prepared by the impregnation method and their catalytic activity for simultaneous hydrolysis and oxidation reactions was investigated in this study. SF6 can be decomposed by catalytic hydrolysis and oxidation. Alumina catalysts have high activity for the decomposition of SF6. However, γ-Al2O3 is deactivated during the hydrolysis and oxidation of SF6, due to the phase transformation of γ-alumina to α-alumina and AlF3. It was confirmed that the stability of the catalyst can be enhanced by using phosphoric acid impregnated over γ-alumina. The impregnation of phosphoric acid over γ-alumina led to the formation of AlPO4 on the surface of the catalyst. The most suitable amount of phosphoric acid on γ-alumina was 4.8wt%. The surface area of the AlPO4/γ-alumina catalyst decreased with increasing amount of phosphoric acid impregnated over γ-alumina and the catalytic activity was also decreased due to the reduction of the surface area. The minimum space velocity should be kept below 5000ml/gcath in order to obtain the maximum efficiency of AlPO4/γ-alumina at 750°C. It was concluded that AlPO4/γ-alumina can be used as a catalyst for the decomposition of SF6 by simultaneous hydrolysis and oxidation.

Bio-hydrogen production from acid hydrolyzed waste ground wheat by dark fermentation

Waste ground wheat was subjected to acid hydrolysis (pH = 3.0) at 90 °C for 15 min using an autoclave. The sugar solution obtained from acid hydrolysis was subjected to dark fermentation for hydrogen gas production after neutralization. In the first set of experiments, initial total sugar concentration was varied between 3.9 and 27.5 g L −1 at constant biomass (cell) concentration of 1.3 g L −1. Biomass concentration was varied between 0.28 g L −1 and 1.38 g L −1 at initial total sugar concentration of 7.2 ± 0.2 g L −1 in the second set of experiments. The highest hydrogen yield (1.46 mol H 2 mol −1 glucose) and the specific formation rate (83.6 ml H 2 g −1 cell h −1) were obtained with 10 g L −1 initial total sugar concentration. Biomass (cell) concentration affected the specific hydrogen production rate yielding the highest rate (1221 ml H 2 g −1 cell h −1) and the yield at the lowest (0.28 g L −1) initial biomass concentration. The most suitable X o / S o ratio, maximizing the yield and specific rate of hydrogen gas formation was X o / S o = 0.037. Dark fermentation of acid hydrolyzed ground wheat was found to be more beneficial as compared to simultaneous bacterial hydrolysis and fermentation.

Kinetic study of a complex triangular reaction system in alkaline aqueous-ethanol medium using on-line transmission FTIR spectroscopy and BTEM analysis

The kinetics of the base-catalyzed reaction of methyl 4-hydroxybenzoate in aqueous-ethanol solvent medium was studied and analyzed via combined on-line transmission FTIR spectroscopy and Band-Target Entropy Minimization (BTEM) technique. This reaction is considered complex since it involves simultaneous hydrolysis and ethanolysis reactions of methyl 4-hydrozybenzoate (MP) to form ethyl 4-hydroxybenzoate (EP) as an intermediate and sodium 4-hydroxybenzoate as a final product. The pure component spectra of the reactive species involved in the reaction were reconstructed using BTEM technique. Their corresponding real concentrations were calculated and subsequently used for analyzing the kinetics of this triangular reaction system. The effects of temperature and solvent mixture compositions were studied. In general, the results show that the rates of both hydrolysis and ethanolysis reactions increase with temperature. Addition of ethanol to the solvent mixture also reduces the rates of the hydrolysis reactions. The effect of solvent mixture on the rate of ethanolysis reaction is more complex and influenced by at least two competing factors, namely the concentration of ethoxide ion in the solution and the stabilization effect on the reactant. The enthalpy and entropy activation parameters, ΔH‡ and ΔS‡, of both the hydrolysis and ethanolysis reactions were determined using the Eyring equation and the activation parameters confirm the associative nature in the elementary steps in these reactions. Finally, it is shown that the dominant synthetic pathway in this triangular system changes from direct hydrolysis of methyl 4-hydrozybenzoate to the indirect pathway via ethanolysis and then hydrolysis depending on the solvent mixture composition.

Coordination Power Adjustment of Surface-Regulating Polymers for Shaping Gold Polyhedral Nanocrystals

PVP (poly(vinyl pyrrolidone)) is a common polymer that behaves as a surface-regulating agent that shapes metal nanocrystals in the polyol process. We have used different polymers containing tertiary amide groups, namely PVCL (poly(vinyl caprolactam)) and PDMAm (poly(N,N-dimethyl acrylamide)), for the synthesis of gold polyhedrons, including octahedrons, cuboctahedrons, cubes, and higher polygons, under the present polyol reaction conditions. The basicity and surface coordination power of the polymers are in the order of PVCL, PVP, and PDMAm. A correlation is observed between the coordination power of the polymers and the resulting gold nanocrystal size. Strong coordination and electron donation from the polymer functional groups to the gold surface restrict particle growth rates, which leads to small nanocrystals. The use of PVCL can yield gold polyhedral structures with small sizes, which cannot be achieved in the reactions with PVP. Simultaneous hydrolysis of the amide group in PDMAm leads to carboxylate functionality, which is very useful for generating chemical and bioconjugates through the formation of ester and amide bonds.

Thermal decomposition mechanisms of MgCl 2·6H 2O and MgCl 2·H 2O

The thermal decomposition mechanisms and the intermediate morphology of MgCl 2·6H 2O and MgCl 2·H 2O were studied using integrated thermal analysis, X-ray diffraction, scanning electron microscope and chemical analysis. The results showed that there were six steps in the thermal decomposition of MgCl 2·6H 2O: producing MgCl 2·4H 2O at 69 °C, MgCl 2·2H 2O at 129 °C, MgCl 2· nH 2O (1 ≤ n ≤ 2) and MgOHCl at 167 °C, the conversion of MgCl 2· nH 2O (1 ≤ n ≤ 2) to Mg(OH)Cl·0.3H 2O by simultaneous dehydration and hydrolysis at 203 °C, the dehydration of Mg(OH)Cl·0.3H 2O to MgOHCl at 235 °C, and finally the direct conversion of MgOHCl to the cylindrical particles of MgO at 415 °C. To restrain the sample hydrolysis and to obtain MgCl 2·H 2O, MgCl 2·6H 2O was first calcined in HCl atmosphere until 203 °C when MgCl 2·H 2O was obtained; HCl gas was then turned off and the calcination process continued, producing Mg 3Cl 2(OH) 4·2H 2O calcined at 203 °C, Mg 3(OH) 4Cl 2 at 220 °C and MgO at 360 °C. The temperature of producing MgO from calcination of MgCl 2·H 2O was lower (360 °C) than that from MgCl 2·6H 2O (415 °C) because of its more reactive intermediate products: the irregular shape and tiny needle-like Mg 3Cl 2(OH) 4·2H 2O particles and the uneven surface porous Mg 3(OH) 4Cl 2 particles. The MgO particles obtained at 360 °C had a flake structure.

The role of acetyl xylan esterase in the solubilization of xylan and enzymatic hydrolysis of wheat straw and giant reed

BackgroundDue to the complexity of lignocellulosic materials, a complete enzymatic hydrolysis into fermentable sugars requires a variety of cellulolytic and xylanolytic enzymes. Addition of xylanases has been shown to significantly improve the performance of cellulases and to increase cellulose hydrolysis by solubilizing xylans in lignocellulosic materials. The goal of this work was to investigate the effect of acetyl xylan esterase (AXE) originating from Trichoderma reesei on xylan solubilization and enzymatic hydrolysis of cellulose.ResultsThe solubilization of xylan in pretreated wheat straw and giant reed (Arundo donax) by xylanolytic enzymes and the impact of the sequential or simultaneous solubilization of xylan on the hydrolysis of cellulose by purified enzymes were investigated. The results showed that the removal of acetyl groups in xylan by AXE increased the accessibility of xylan to xylanase and improved the hydrolysis of xylan in pretreated wheat straw and giant reed. Solubilization of xylan led to an increased accessibility of cellulose to cellulases and thereby increased the hydrolysis extent of cellulose. A clear synergistic effect between cellulases and xylanolytic enzymes was observed. The highest hydrolysis yield of cellulose was obtained with a simultaneous use of cellulases, xylanase and AXE, indicating the presence of acetylated xylan within the cellulose matrix. Acetylated xylobiose and acetylated xylotriose were produced from xylan without AXE, as confirmed by atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry.ConclusionsThe results in this paper demonstrate that supplementation of xylanase with AXE enhances the solubilization of xylan to some extent and, consequently, increases the subsequent hydrolysis of cellulose. The highest hydrolysis yield was, however, obtained by simultaneous hydrolysis of xylan and cellulose, indicating a layered structure of cellulose and xylan chains in the cell wall substrate. AXE has an important role in the hydrolysis of lignocellulosic materials containing acetylated xylan.

Effects of reaction conditions on the acid-catalyzed hydrolysis of miscanthus dissolved in an ionic liquid

Experiments were conducted to study the effects of reaction conditions on the hydrolysis of miscanthus dissolved in 1-ethyl-3-methylimidazolium chloride ([Emim][Cl]) catalyzed by H2SO4. It was determined that while there is a small co-inhibition effect associated with the simultaneous hydrolysis of the cellulosic and hemicellulosic portions of miscanthus, the largest rate decreases were observed for the hydrolysis of the hemicellulosic portion. This rate decrease was attributed to the chemical linkage between hemicellulose and lignin in miscanthus, which could be broken with chemical pretreatment. While chemical pretreatment increased the rate of the hydrolysis of the hemicellulosic component, delignification showed no further benefit. The rate of hydrolysis was determined to be first order in concentrations of β-1,4 glycosidic linkage and acid, and zero order in water concentration. The activation energy for the hydrolysis of the glycosidic linkages in the cellulosic and hemicellulosic components were determined to be 95 kJ mol−1 and 114 kJ mol−1 respectively. Progressive addition of water during the first hour of the reaction increased conversion and selectivity to saccharine products, while limiting dehydration of the sugars formed. The conversion of the cellulosic portion of miscanthus could be increased after the first hour of the reaction by increasing the reactor temperature. While miscanthus is only partially soluble in [Emim][Cl], it was found that the initial miscanthus loading could be increased to 9 wt% before significant yield decreases attributed to solubility limitations of the cellulosic component were observed. By proper adjustment of reaction conditions, it was possible to achieve yields of sugars approaching 84% from the cellulosic and hemicellulosic components of miscanthus, with minimal dehydration of the sugars to furans.

Significant thermal energy reduction in lactic acid production process

Lactic acid is widely used as a raw material for the production of biodegradable polymers and in food, chemical and pharmaceutical industries. The global market for lactic acid is expected to reach 259 thousand metric tons by the year 2012. For batch production of lactic acid, the traditional process includes the following steps: (i) esterification of impure lactic acid with methanol in a batch reactor to obtain methyl lactate (ester), (ii) separation of the ester in a batch distillation, (iii) hydrolysis of the ester with water in a batch reactor to produce lactic acid and (iv) separation of lactic acid (in high purity) in a batch distillation. Batch reactive distillation combines the benefit of both batch reactor and batch distillation and enhances conversion and productivity (Taylor and Krishna, 2000 [1]; Mujtaba and Macchietto, 1997 [2]). Therefore, the first and the last two steps of the lactic acid production process can be combined together in batch reactive distillation (Fig. 1) processes. However, distillation (batch or continuous) is an energy intensive process and consumes large amount of thermal energy (via steam). This paper highlights how significant (over 50%) reduction in thermal energy consumption can be achieved for lactic acid production process by carefully controlling the reflux ratio but without compromising the product specification. In this paper, only the simultaneous hydrolysis of methyl lactate ester and the separation of lactic acid using batch reactive distillation is considered.

Microwave-assisted hydrolysis and extraction of tricyclic antidepressants from human hair

The objective of this research was to develop, optimize, and validate a modern, rapid method of preparation of human hair samples, using microwave irradiation, for analysis of eight tricyclic antidepressants (TCADs): nordoxepin, nortriptyline, imipramine, amitriptyline, doxepin, desipramine, clomipramine, and norclomipramine. It was based on simultaneous alkaline hair microwave-assisted hydrolysis and microwave-assisted extraction (MAH–MAE). Extracts were analyzed by high-performance liquid chromatography with diode-array detection (HPLC–DAD). A mixture of n-hexane and isoamyl alcohol (99:1, v/v) was used as extraction solvent and the process was performed at 60°C. Application of 1.0 mol L−1 NaOH and microwave irradiation for 40 min were found to be optimum for hair samples. Limits of detection ranged from 0.3 to 1.2 μg g−1 and LOQ from 0.9 to 4.0 μg g−1 for the different drugs. This enabled us to quantify them in hair samples within average therapeutic concentration ranges.

Highly efficient production of d-lactate by Sporolactobacillus sp. CASD with simultaneous enzymatic hydrolysis of peanut meal

Highly efficient D-lactate production by Sporolactobacillus sp. strain CASD was demonstrated in this study. Peanut meal was found to be a better nutrient than yeast extract, soybean meal, soybean peptone, corn steep, liquor beef extract, and ammonium sulfate in the production of D-lactate. To improve the utilization of peanut meal, the material was enzymatically hydrolyzed and simultaneously utilized as the nitrogen source in D-lactate fermentation. Very high D-lactate production (207 g/L) was obtained using 40 g/L of peanut meal in 30-L fed-batch fermentation, with the average productivity of 3.8 g/(L·h) and optical purity of 99.3%. The production of such a high concentration of optically pure D-lactate by strain CASD, with the simultaneous enzymatic hydrolysis of peanut meal and fermentation, represents a new cost-efficient and integrated method for D-lactate production using agricultural by-products.

Simultaneous pressurized enzymatic hydrolysis extraction and clean up for arsenic speciation in seafood samples before high performance liquid chromatography–inductively coupled plasma-mass spectrometry determination

The feasibility of pressurized conditions to assist enzymatic hydrolysis of seafood tissues for arsenic speciation was novelty studied. A simultaneous in situ (in cell) clean-up procedure was also optimized, which speeds up the whole sample treatment. Arsenic species (As(III), MMA, DMA, As(V), AsB and AsC) were released from dried seafood tissues using pepsin as a protease, and the arsenic species were separated/quantified by anion exchange high performance liquid chromatography (HPLC) coupled to inductively coupled plasma-mass spectrometry (ICP-MS). Variables inherent to the enzymatic activity (pH, temperature and ionic strength), the amount of enzyme (pepsin), and factors affecting pressurization (pressure, static time, number of cycles and amount of dispersing agent, C-18) were fully evaluated. Pressurized assisted enzymatic hydrolysis (PAEH) with pepsin can be finished after few minutes (two cycles of 2 min each one plus 3 min to reach the hydrolysis temperature of 50 °C). A total sample solubilisation is not achieved after the procedure, however it is efficient enough for breaking down certain bonds of bio-molecules and for releasing arsenic species. The developed method has been found to be precise (RSDs lower than 6% for As(III), DMA and As(V); and 3% for AsB) and sensitive (LOQs of 18.1, 36.2, 35.7, 28.6, 20.6 and 22.5 ng/g for As(III), MMA, DMA, As(V), AsB and AsC, respectively). The optimized methodology was successfully applied to different certified reference materials (DORM-2 and BCR 627) which offer certified AsB and DMA contents, and also to different seafood products (mollusks, white fishes and cold water fishes).

Improving the fermentability of enzymatic hydrolysates of lignocellulose through chemical in-situ detoxification with reducing agents

Inhibitory lignocellulose hydrolysates were treated with the reducing agents dithionite and sulfite to achieve improved fermentability. Addition of these reducing agents (in the concentration range 5.0-17.5 mM) to enzymatic hydrolysates of spruce wood or sugarcane bagasse improved processes based on both SHF (simultaneous hydrolysis and fermentation) and SSF (simultaneous saccharification and fermentation). The approach was exemplified in ethanolic fermentations with Saccharomyces cerevisiae and by using hydrolysates with sugar concentrations>100 g/L (for SHF) and with 10% dry-matter content (for SSF). In the SHF experiments, treatments with dithionite raised the ethanol productivities of the spruce hydrolysate from 0.2 to 2.5 g×L(-1)×h(-1) and of the bagasse hydrolysate from 0.9 to 3.9 g×L(-1)×h(-1), values even higher than those of fermentations with reference sugar solutions without inhibitors. Benefits of the approach include that the addition of the reducing agent can be made in-situ directly in the fermentation vessel, that the treatment can be performed at a temperature and pH suitable for fermentation, and that the treatment results in dramatically improved fermentability without degradation of fermentable sugars. The many benefits and the simplicity of the approach offer a new way to achieve more efficient manufacture of fermentation products from lignocellulose hydrolysates.