Fe-MOF/AuNP-based ratiometric electrochemical immunosensor for the detection of deoxynivalenol in grain products.

A ratiometric assay was designed to improve the sensitivity and reliability of electrochemical immunosensors for deoxynivalenol (DON) detection. The indicator signal caused by the Fe-based metal-organic framework nanocomposites loaded with gold nanoparticles and the internal reference signal from the [Fe(CN)6]3-/4- in the electrolyte came together at the immunosensor. When immunoreactivity occurred, the indicator signals decreased as the concentration of DON increased, while the internal reference signals increased slightly. The ratio of the indicator signal to the internal reference signal was available for reproducible and sensitive monitoring of DON. The prepared immunosensor showed excellent performance in the range from 0.5 to 5000 pg mL-1, and the detection limit was 0.0166 pg mL-1. The immunosensor achieved satisfactory detection toward DON in spiked and actual samples and has a promising application in the control of DON in grain products.

Engineering Corynebacterium glutamicum Mutants for 3-Methyl-1-butanol Production.

3-Methyl-1-butanol (3MB) is a promising biofuel that can be produced from 2-ketoisocaproate via the common L-leucine biosynthesis pathway. Corynebacterium glutamicum was chosen as a host bacterium because of its strong resistance to isobutanol. In the current study, several strategies were designed to overproduce 3MB in C. glutamicum through a non-fermentation pathway. The engineered C. glutamicum mutant was obtained by silencing the pyruvate dehydrogenase gene complex (aceE) and deleting the lactic dehydrogenase gene (ldh), followed by mutagenesis with diethyl sulfate (DES) and selection with Fmoc-3-4-thiazolyl-L-alanine (FTA). The mutant could produce 659 mg/L of 3MB after 12 h of incubation. To facilitate carbon flux to 3MB biosynthesis, the engineered recombinant was also constructed without branched-chain acid aminotransferase (ilvE) activity by deleting the ilvE gene. This recombinant could produce 697 mg/L of 3MB after 12 h of incubation.

One-step hydroprocessing of fatty acids into renewable aromatic hydrocarbons over Ni/HZSM-5: insights into the major reaction pathways.

For high caloricity and stability in bio-aviation fuels, a certain content of aromatic hydrocarbons (AHCs, 8-25 wt%) is crucial. Fatty acids, obtained from waste or inedible oils, are a renewable and economic feedstock for AHC production. Considerable amounts of AHCs, up to 64.61 wt%, were produced through the one-step hydroprocessing of fatty acids over Ni/HZSM-5 catalysts. Hydrogenation, hydrocracking, and aromatization constituted the principal AHC formation processes. At a lower temperature, fatty acids were first hydrosaturated and then hydrodeoxygenated at metal sites to form long-chain hydrocarbons. Alternatively, the unsaturated fatty acids could be directly deoxygenated at acid sites without first being saturated. The long-chain hydrocarbons were cracked into gases such as ethane, propane, and C6-C8 olefins over the catalysts' Brønsted acid sites; these underwent Diels-Alder reactions on the catalysts' Lewis acid sites to form AHCs. C6-C8 olefins were determined as critical intermediates for AHC formation. As the Ni content in the catalyst increased, the Brønsted-acid site density was reduced due to coverage by the metal nanoparticles. Good performance was achieved with a loading of 10 wt% Ni, where the Ni nanoparticles exhibited a polyhedral morphology which exposed more active sites for aromatization.

Combined use of GAP and AOX1 promoters and optimization of culture conditions to enhance expression of Rhizomucor miehei lipase.

Rhizo mucor miehei lipase (RML) is an industrially important enzyme, but its application is limited due to its high cost. In this study, a series of measures such as codon optimization, propeptide addition, combined use of GAP and AOX1 promoters, and optimization of culture conditions were employed to increase the expression of RML. Three transformants of the constitutive-inducible combined Pichia pastoris strains were generated by transforming the pGAPZαA-rml vector into the pPIC9K-rml/GS115 strain, which resulted in high-expression yields of RML. Using the shake flask method, highest enzyme activity corresponding to 140 U/mL was observed in the strain 3-17, which was about sixfold higher than that of pPIC9K-rml/GS115 or pGAPZαA-rml/GS115. After optimization of culture conditions by response surface methodology, the lipolytic activity of strain 3-17 reached 175 U/mL in shake flasks. An increase in the copy number simultaneously with the synergistic effect provided by two promoters led to enhanced degree of protein expression.

Production of C4 and C5 branched-chain alcohols by engineered Escherichia. coli.

Higher alcohols, longer chain alcohols, contain more than 3 carbon atoms, showed close energy advantages as gasoline, and were considered as the next generation substitution for chemical fuels. Higher alcohol biosynthesis by native microorganisms mainly needs gene expression of heterologous keto acid decarboxylase and alcohol dehydrogenases. In the present study, branched-chain α-keto acid decarboxylase gene from Lactococcus lactis subsp. lactis CICC 6246 (Kivd) and alcohol dehydrogenases gene from Zymomonas mobilis CICC 41465 (AdhB) were transformed into Escherichia coli for higher alcohol production. SDS-PAGE results showed these two proteins were expressed in the recombinant strains. The resulting strain was incubated in LB medium at 37 °C in Erlenmeyer flasks and much more 3-methyl-1-butanol (104 mg/L) than isobutanol (24 mg/L) was produced. However, in 5 g/L glucose-containing medium, the production of two alcohols was similar, 156 and 161 mg/L for C4 (isobutanol) and C5 (3-methyl-1-butanol) alcohol, respectively. Effects of fermentation factors including temperature, glucose content, and α-keto acid on alcohol production were also investigated. The increase of glucose content and the adding of α-keto acids facilitated the production of C4 and C5 alcohols. The enzyme activities of pure Kivd on α-ketoisovalerate and α-ketoisocaproate were 26.77 and 21.24 µmol min(-1) mg(-1), respectively. Due to its ability on decarboxylation of α-ketoisovalerate and α-ketoisocaproate, the recombinant E. coli strain showed potential application on isoamyl alcohol and isobutanol production.

Water-saving analysis on an effective water reuse system in biodiesel feedstock production based on Chlorella zofingiensis fed-batch cultivation.

The micralgae-based biofuel obtained from dairy wastewater (DWW) is considered a promising source of energy. However, this process consumes water due to the concentration of wastewater being normally too high for some micoralgae cultivation, and dilution is always needed. In this work, the cultivation of microalgae has been examined in non-recirculated water (NR) and recirculated water systems (R). The growth of Chlorella zofingiensis and the nutrient removal of DWW have been recorded. The comparison indicates the R had a little more advantage in biomass and lipid output (1.55, 0.22 g, respectively) than the NR (1.51, 0.20 g, respectively). However, the total chemical oxygen demand (COD), total Kjeldahl nitrogen (TKN), and total phosphorus (TP) removals of the R were lower than those of the NR system during the culture. The highest removal of total COD, TKN, and TP were 85.05%, 93.64%, and 98.45%, respectively. Furthermore, no significant difference has been observed in the higher heating value and lipid content of the biomass of the R and NR. The results show the R can save 30% of the total water input during the culture. All above results indicate the R system has great potential in industry.

Factors affecting the performance of a single-chamber microbial fuel cell-type biological oxygen demand sensor.

Microbial fuel cells (MFCs) are devices that exploit microorganisms as biocatalysts to degrade organic matter or sludge present in wastewater (WW), and thereby generate electricity. We developed a simple, low-cost single-chamber microbial fuel cell (SCMFC)-type biochemical oxygen demand (BOD) sensor using carbon felt (anode) and activated sludge, and demonstrated its feasibility in the construction of a real-time BOD measurement system. Further, the effects of anodic pH and organic concentration on SCMFC performance were examined, and the correlation between BOD concentration and its response time was analyzed. Our results demonstrated that the SCMFC exhibited a stable voltage after 132 min following the addition of synthetic WW (BOD concentration: 200 mg/L). Notably, the response signal increased with an increase in BOD concentration (range: 5-200 mg/L) and was found to be directly proportional to the substrate concentration. However, at higher BOD concentrations (>120 mg/L) the response signal remained unaltered. Furthermore, we optimized the SCMFC using synthetic WW, and tested it with real WW. Upon feeding real WW, the BOD values exhibited a standard deviation from 2.08 to 8.3% when compared to the standard BOD5 method, thus demonstrating the practical applicability of the developed system to real treatment effluents.

Continuous anaerobic digestion of food waste and design of digester with lipid removal.

Separation of municipal solid waste has been implemented in many cities in China. As a major component of municipal solid waste, food waste can be treated by anaerobic digestion (AD) for energy production. To provide reference data for disposing of food waste through engineering applications, continuous AD was carried out under various organic loading rates (OLRs) at 27 +/- 2 degrees C in the laboratory. The anaerobic reactor was stable with pH 7.0-7.1, total volatile fatty acid (VFA) concentrations of 206-746 mg/L, and NH4+ -N concentrations of 525-1293 mg/L when the OLR was 1.118-5.588 kg volatile solids (VS)/m(3) x d. The maximum volumetric biogas production rate was 4.41 L/L x d when the OLR was increased to 5.588 kg VS/m(3) x d with a hydraulic retention time of 30 d. When the OLR was increased to 6.706 and 8.382 kg VS/m(3) x d, biogas production was seriously inhibited by VFAs, with maximum total VFA and propionate concentrations of 8738 mg/L and 2864 mg/L, respectively. Due to the incomplete degradation of lipids, the specific methane production rate of 353-488 L/kg VS accounted for 55.2-76.3% of the theoretical methane potential calculated based on the component composition. A retrofitted anaerobic digester with lipid removal was designed to improve the efficiency.

[Bioconversion of C1 gases and genetic engineering modification of gas-utilizing microorganisms].

C1 gases including CO, CO2 and CH4, are mainly derived from terrestrial biological activities, industrial waste gas and gasification syngas. Particularly, CO2 and CH4 are two of the most important greenhouse gases contributing to climate change. Bioconversion of C1 gases is not only a promising solution to addressing the problem of waste gases emission, but also a novel route to produce fuels or chemicals. In the past few years, C1-gas-utilizing microorganisms have drawn much attention and a variety of gene-editing technologies have been applied to improve their product yields or to expand product portfolios. This article reviewed the biological characteristics, aerobic or anaerobic metabolic pathways as well as the metabolic products of methanotrophs, autotrophic acetogens, and carboxydotrophic bacteria. In addition, gene-editing technologies (e.g. gene interruption technology using homologous recombination, group Ⅱ intron ClosTron technology, CRISPR/Cas gene editing and phage recombinase-mediated efficient integration of large DNA fragments) and their application in these C1-gas-utilizing microorganisms were also summarized.

An electrochemical immunosensor based on prussian blue@zeolitic imidazolate framework-8 nanocomposites probe for the detection of deoxynivalenol in grain products.

The presence of deoxynivalenol (DON) in grains poses a threat to human health, which is critical for sensitive detection of DON. In this electrochemical immunosensor, zeolitic imidazolate framework-8 (ZIF-8) loaded with Prussian blue (PB) nanoparticles was coated by polydopamine (PDA) as a redox probe. The high porosity of ZIF-8, the unique electrochemical activity of PB and the outstanding electrical conductivity of PDA improved the sensitivity of the immunosensor. Under the optimized conditions, the peak current in differential pulse voltammetry displayed a good linear relationship over DON concentrations in a range of 0.1-5000 pg mL-1 with a detection limit of 0.0186 pg mL-1. In addition, the immunosensor also had good selectivity and stability. Good recoveries of 85.67 to 118.00 % have been achieved for the detection of DON in spiked grain products. This new strategy exhibits great potential for simple and rapid detection of DON in grain and feed products.

Artificial intelligence techniques to optimize the EDC/NHS-mediated immobilization of cellulase on Eudragit L-100.

Two artificial intelligence techniques, namely artificial neural network (ANN) and genetic algorithm (GA) were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) concentration, N-hydroxysuccinimide (NHS) concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the central composite design were used to train ANN by back-propagation algorithm, and the result showed that the trained ANN fitted the data accurately (correlation coefficient R(2) = 0.99). Then a maximum immobilization efficiency of 88.76% was searched by genetic algorithm at a EDC concentration of 0.44%, NHS concentration of 0.37% and a coupling time of 2.22 h, where the experimental value was 87.97 ± 6.45%. The application of ANN based optimization by GA is quite successful.

Facile synthesis of lignin-based epoxy resins with excellent thermal-mechanical performance.

Up to now, various approaches have been used to fabricate lignin-based epoxy thermosets by utilizing lignin or lignin-derivatives, but there is still lack of a simple, effective and environmental-friendly pathway for producing lignin-based epoxy resins from industrial lignin. In this work, a novel strategy - one-pot to synthesize phenolated lignin incorporated novolac epoxy networks (PLIENs) was proposed. As expected, PLIENs obtained from the novel route exhibited preferable mechanical and thermal properties compared with the epoxy resins which obtained from common route. Moreover, increasing the loading of lignin did not significantly deteriorate the thermal-mechanical performance of cured epoxy resins. However, the Tg of PLIENs was slightly lowered compared with conventional petroleum-based epoxy resins (DGEBA). Nonetheless, the flexural strength and storage modulus of PLIENs were higher than that of DGEBA. Especially, the char yield of PLIENs at 800 °C was up to 28.9%, much higher than that of DGEBA (only 6.9%), which indicated that lignin has a certain promoting effect on the flame retardancy of epoxy resins. This research provides a new insight for producing commercially viable lignin-based epoxy thermosets.

The effect of enzyme loading, alcohol/acid ratio and temperature on the enzymatic esterification of levulinic acid with methanol for methyl levulinate production: a kinetic study.

As an important bio-based chemical, methyl levulinate (ML) can be produced via enzymatic esterification of levulinic acid with methanol. A kinetic model is developed in this work based on the law of mass action and reaction reversibility, to investigate the effect of enzyme loading, alcohol/acid ratio and temperature on ML yield. Data analysis shows that newly developed binary regression is apparently more persuasive than the commonly used unitary regression. Kinetic study reveals: (1) rate constants of esterification/hydrolysis increase with increasing enzyme loading, while their ratio (equilibrium constant) remains invariant. (2) Methanol has no toxicity towards lipase, and hence, neither the rate constants of esterification/hydrolysis nor the equilibrium constant are affected by alcohol/acid ratio. (3) Both rate constants of esterification/hydrolysis and the equilibrium constant increase with temperature elevation, and their relationships agree with Arrhenius equation and Van't Hoff equation, respectively. (4) The esterification is endothermic and spontaneous. In total, the application of binary regression analysis for the developed model to study the enzymatic esterification kinetics is quite successful.

A one-step deconstruction-separation organosolv fractionation of lignocellulosic biomass using acetone/phenoxyethanol/water ternary solvent system.

A novel ternary solvent system for organosolv fractionation of lignocellulosic biomass, named APW process, which is composed of acetone, phenoxyethanol and water with the advantages of monophasic deconstruction and biphasic separation of components was developed. Through fractionation of amorpha as a case study, a monophasic APW solution (acetone/phenoxyethanol/water = 5:11:4, volume ratio) with the best lignin affinity was constructed based on Hansen solubility parameters. According to Taguchi experimental design, the optimal conditions were 130 °C, 70 min, 0.15 M sulfuric acid and 20 LSR. Under optimal conditions, removal of lignin and hemicellulose reached 95.60% and 98.39%, respectively. While 80.48% of cellulose was retained in residue and its digestibility was 80.36%. Then, 83.74% of hemicellulose was recovered from aqueous as sugars, and 35.64% of lignin was recovered by precipitation. Moreover, APW process also have effective fractionation of sugarcane bagasse, corn cob and pine, cellulose and hemicellulose recovery were both over 80%.

Impact of Alkaline Pretreatment Condition on Enzymatic Hydrolysis of Sugarcane Bagasse and Pretreatment Cost.

A combined severity factor (RCSF) which is usually used to evaluate the effectiveness of hydrothermal pretreatment at above 100 °C had been developed to assess the influence of temperature, time, and alkali loading on pretreatment and enzymatic hydrolysis of lignocellulose. It is not suitable for evaluating alkaline pretreatment effectiveness at lower than 100 °C. According to the reported deducing process, this study modified the expression of [Formula: see text] as [Formula: see text] which is easier and more reasonable to assess the effectiveness of alkaline pretreatment. It showed that RCSF exhibited linear trend with lignin removal, and quadratic curve relation with enzymatic hydrolysis efficiency (EHE) at the same temperature. The EHE of alkali-treated SCB could attain the maximum value at lower RCSF, which indicated that it was not necessary to continuously enhance strength of alkaline pretreatment for improving EHE. Within a certain temperature range, the alkali loading was more important than temperature and time to influence pretreatment effectiveness and EHE. Furthermore, the contribution of temperature, time, and alkali loading to pretreatment cost which was seldom concerned was investigated in this work. The alkali loading contributed more than 70% to the pretreatment cost. This study laid the foundation of further optimizing alkaline pretreatment to reduce cost for its practical application.

Synthesis of a Stable Solid Acid Catalyst from Chloromethyl Polystyrene through a Simple Sulfonation for Pretreatment of Lignocellulose in Aqueous Solution.

A stable solid acid catalyst, SCPR140-1, was synthesized from chloromethyl polystyrene resin (CPR) and used for catalytic pretreatment of corncob in aqueous solution. Under the optimized pretreatment condition, 73.07 % of xylose was directly obtained, and the enzymatic digestibility of treated residue reached up to 94.65 %, indicating that the SCPR140-1 had high selectivity for xylose production and effectively deconstructed the structure of corncob. The -CH2 Cl group of CPR was substituted by -SO3 H through the sulfonation, and the -SO3 H was stably bound on the catalyst during the pretreatment process. Compared with other similar reports, the SCPR140-1 was not only synthesized through a simpler process but also had a more stable catalytic activity during multiple recycling runs.

Enhanced methane production from anaerobic digestion of hybrid Pennisetum by selectively removing lignin with sodium chlorite.

To enhance the biodegradability and methane production of hybrid Pennisetum, a pretreatment method with high selectivity for lignin removal, namely sodium chlorite/acetic acid (SCA) pretreatment, was examined in this work. Results showed that SCA pretreatment can selectively remove lignin with minimal impact on cellulose and hemicellulose. After up to 200 min of SCA treatment, 79.4% of lignin was removed and over 90% of the holocellulose was retained. The physicochemical changes after pretreatment were analyzed by confocal laser scanning microscopy, X-ray diffractometer and Fourier transform infrared spectroscopy, showing that the majority of lignin was removed from secondary cell walls and cell middle lamella while the chlorite-resistant lignin remained in the cell corner. Lignin removal significantly enhanced the biodegradability from 59.6% to 86.4% and increased methane production by 38.3%. Energy balance showed that SCA pretreatment was efficient to increase the energy output of hybrid Pennisetum.

Effects of fermentative and non-fermentative additives on silage quality and anaerobic digestion performance of Pennisetum purpureum.

The effect of additives on the silage quality, microbial community, and anaerobic digestion performance of Pennisetum purpureum with high moisture content was studied. The sample treated with a mixed additive had best silage quality with the lowest pH and highest lactic acid/acetic acid ratio. Different additives influenced the dominant desirable bacteria. Correspondingly, Enterobacter was the dominant bacterial genus for sample with non-fermentative additives, whereas for the samples with fermentative or mixed additives, both Enterobacter and Lactobacillus had high relative abundance. The parameters of NH3-N, hemicellulose and lactic acid were positively correlated with the specific methane yield, while the lignin content was inversely correlated with the specific methane yield. The higher specific methane yield of 293.81 ± 0.15-334.69 ± 22.75 mL/g VS was obtained for samples treated with fermentative additive. Therefore, the mixed additive and fermentative additive are recommended for the silage of material with high-moisture content to improve the silage quality and methane yield.

Depolymerization of holocellulose from Chinese herb residues by the mixture of lignin-derived deep eutectic solvent with water.

The depolymerization of biomass carbohydrate polymers usually happened in homogeneous medium, just a few in heterogeneous solution. Herein, holocellulose from two Chinese herb residues Cortex albiziae (HRCA) and Heteropogon contortus (HRHC) was prepared and characterized. Deep eutectic solvent (DES) of choline chloride/p-coumaric acid collaborating with water was employed for the dissociation of those two holocelluloses and selectively dissolved hemicellulose into xylose with maximal yields of 81.50 % and 72.47 %, respectively. Most cellulose remained as the polymer state with a maximum solubility of 9.38 %. The synergistic action of DES and water was investigated to unveil the depolymerization mechanism and the roles of each component in depolymerizing procedure. It was water that released hemicellulose polymers and dissolved polymers to soluble short oligosaccharides, and DES rapidly cleaved the latter to xylose. Contrarily, DES broke cellulose to only soluble cello-oligosaccharides, but water exhibited the ability to hydrolyze cellulose to glucose.

Effect of a Nonionic Surfactant on Enzymatic Hydrolysis of Lignocellulose Based on Lignocellulosic Features and Enzyme Adsorption.

Reduction in the adsorption of cellulase onto lignin has been thought to be the common reason for the improvement of enzymatic hydrolysis of lignocellulose (EHLC) by a nonionic surfactant (NIS). Few research studies have focused on the relationship between lignocellulosic features and NIS for improving EHLC. This study investigated the impact of Tween20 on the enzymatic hydrolysis and enzyme adsorption of acid-treated and alkali-treated sugarcane bagasse (SCB), cypress, and Pterocarpus soyauxii (PS) with and without being ground. After addition of Tween20, the adsorption of cellulase onto unground and ground alkali-treated SCB increased, and the unground acid-treated SCB exhibited little change in adsorption cellulase, while other unground and ground, treated samples showed decreased cellulase adsorption. Tween20 could improve the enzymatic hydrolysis of acid-treated SCB, while it had little influence on the enzymatic hydrolysis of other treated materials. After being ground, both cellulase adsorption and enzymatic hydrolysis of treated lignocelluloses increased, and Tween20 could enhance the enzymatic hydrolysis of acid-treated materials while hardly affected the enzymatic hydrolysis of alkali-treated materials. This indicated that the promotion effect of Tween20 on enzymatic hydrolysis of treated lignocellulose could not be mainly ascribed to the hindrance of Tween20 to cellulase adsorption on lignin but was related to the lignocellulosic features such as hemicellulose removal and surface morphology changes.