Theoretical study on electrocatalytic carbon dioxide reduction over copper with copper-based layered double hydroxides.

The conversion of CO2 into valuable fuels and multi-carbon chemical substances by electrical energy is an effective strategy to solve environmental problems by using renewable energy sources. In this work, the density functional theory (DFT) method is used to reveal the electrocatalytic mechanism of CO2 reduction reaction (CO2RR) over the surface of CuAl-Cl-layered double hydroxides (LDHs) with Cu monoatoms (Cu@CuAl-Cl-LDH), Cu2 diatoms (Cu2@CuAl-Cl-LDH), orthotetrahedral Cu4 clusters (Td-Cu4@CuAl-Cl-LDH) and planar Cu4 clusters (Pl-Cu4@CuAl-Cl-LDH). The active sites, density of states, adsorption energy, charge density difference and free energy are calculated. The results show that CO2RR over all the above five catalysts can generate C2 products. Pl-Cu4@CuAl-Cl-LDH tends to generate C2H5OH, while the remaining four structures all tend to produce C2H4. Cuδ+ favors CO2RR, and Td-Cu4@CuAl-Cl-LDH with a larger positively charged area at the active site has the better electrocatalytic performance among the calculated systems with a maximum step height of 0.78 eV. The selectivity of the products C2H4 and C2H5OH depends on the dehydration of the intermediate *C2H2O to *C2H3O or *CCH; if the dehydration produces *CCH intermediate, the final product is C2H4, and if no dehydration occurs, C2H5OH is produced. This work provides theoretical information and guidance for further rational design of efficient CO2RR catalysts for energy saving and emission reduction.

Succession of sulfur bacteria during decomposition of cyanobacterial bloom biomass in the shallow Lake Nanhu: An ex situ mesocosm study.

Previous studies of the dynamics of sulfate-reducing bacteria (SRB) and sulfur-oxidizing bacteria (SOB) have focused on deep stratified lakes. The objective of this study is to present an in-depth investigation of the structure and dynamics of sulfur bacteria (including SRB and SOB) in the water column of shallow freshwater lakes. A cyanobacterial bloom biomass (CBB)-amended mesocosm experiment was conducted in this study, in which water was taken from a shallow eutrophic lake with sulfate levels near 40 mg L-1. Illumina sequencing was used to investigate SRB and SOB species involved in CBB decomposition and the effects of the increases in sulfate input on the water column microbial community structure. The accumulation of dissolved sulfide (∑H2S) produced by SRB during CBB decomposition stimulated the growth of SOB, and ∑H2S was then oxidized back to sulfate by SOB in the water column. Chlorobaculum sequences (the main SOB species in the study) were significantly influenced by increases in sulfate input, with relative abundance increasing approximately four-fold in treatments amended with 40 mg L-1 sulfate (referred to as 40S) when compared to the treatment without additional sulfate addition (referred to as CU). Additionally, an increase in SOB number was observed from day 26-37, concurrent with the decrease in SRB number, indicating the succession of sulfur bacteria. These findings suggest that biological sulfur oxidation and succession of sulfur bacteria occur in the water column during CBB decomposition in shallow freshwater ecosystems, and the increases in sulfate input stimulate microbial sulfur oxidation.

Construction of a trifunctional cellulase and expression in Saccharomyces cerevisiae using a fusion protein.

BACKGROUND: Cellulose is the most important component of lignocellulose, and its degradation requires three different types of enzymes to act synergistically. There have been reports of single gene duality, but no gene has been described to have more than two functions. Cloning and expression of fusion cellulases containing more than two kinds of catalytic domains has not been reported thus far. RESULTS: We synthesized three different cellulase genes and linked the three catalytic domains with a (G4S)3 flexible linker. The trifunctional cellulase gene (BCE) containing three types of cellulase functions was constructed and expressed in S. cerevisiae successfully. The ß-glucosidase, the exoglucanase and the endoglucanase activity of the trifunctional cellulase BCE reached 16.80 IU/mg, 2.26 IU/mg and 20.67 IU/mg, which was 46.27, 6.73 and 46.20% higher than the activities of the ß-glucosidase BG, the endoglucanase CBH and the endoglucanase EG. The filter paper enzyme activity of BCE was higher than those of BG, CBH and EG, reached 2.04 IU/mg. CONCLUSIONS: The trifunctional cellulase BCE was designed based on ß-glucosidase BG, endoglucanase EG and exoglucanase CBH, and it possessed ß-glucosidase activity, endoglucanase activity and exoglucanase activity simultaneously. The BCE has better filter paper activity, it means the potential practical application.

[Screen astragalosides from Huangqi injections by LC-TOF-MS-based mass defect filtering approach].

The samples of Huangqi injection (HI) were analyzed by liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (LC-TOF-MS), and both positive and negative ion modes were employed to obtain the LC-TOF-MS analysis information of chemical compounds in HI. Then the mass defect filtering (MDF) approach, which was developed based on the previously published articles, was utilized to rapidly screen the astragalosides from the obtained LC-TOF-MS data. Each screened astragaloside was confirmed by the presence of no less than 2 quasi-molecular ions. All the screened astragalosides were then tentatively assigned according to the parent ion and daughter ion information. Finally, a total of 62 astragalosides were screened and characterized from the HI samples, including 15 new detected ones. The identification results indicated that acetylation, hydrogenation, dehydrogenation, methoxylation and hydration might be the major conversion reactions involved in the formation of the astragalosides. The LC-TOF-MS-based MDF approach was proved to be a feasible and efficient tool to screen the chemical constituents in complex matrices such as herbal medicines.

Urinary metabolomics reveals the therapeutic effect of HuangQi Injections in cisplatin-induced nephrotoxic rats.

The side effects of cisplatin (CDDP), notably nephrotoxicity, greatly limited its use in clinical chemotherapy. HuangQi Injections (HI), a commonly used preparation of the well-known Chinese herbal medicine Astragali radix, appeared to be promising treatment for nephrotoxicity without compromising the anti-tumor activity of CDDP. In this study, the urinary metabolomics approach using liquid chromatography time of flight mass spectrometry (LC-TOF/MS) was developed to assess the toxicity-attenuation effects and corresponding mechanisms of HI on CDDP-exposed rats. As a result, successive administration of HI significantly recovered the decline of body weight and downregulated the abnormal increase of serum creatinine and urea. HI partly restored the CDDP-induced alteration of metabolic profiling back into normal condition. Totally 43 toxicity-attenuation potential biomarkers were screened and tentatively identified, which were involved in important metabolic pathways such as amino acid metabolism, TCA cycle, fatty acid metabolism, vitamin B6 metabolism and purine metabolism. The results clearly revealed that HI could alleviate CDDP-induced nephrotoxicity and improve the disturbed metabolic balance induced by repeated CDDP exposure. The present study provided reliable evidence for the protective effect of HI on CDDP-induced toxicity with the multi-target pharmacological characteristics.

Construction of recombinant Yarrowia lipolytica and its application in bio-transformation of lignocellulose.

Lignocellulose is a polysaccharide and an abundant biomass resource that widely exists in grains, beans, rice, and their by-products. Over 10 million tons of lignocellulose resources and processing products are produced every year in China. Three recombinant Y. lipolytica strains with cellulase (ß-glucosidase, endoglucanase and cellobiohydrolase) were constructed. The enzymatic activities of these enzymes were 14.181 U/mL, 16.307 U/mL, and 17.391 U/mL, respectively. The whole cell cellulases were used for a stover bio-transformation. The celluloses in the stover were partly degraded by the cellulases, and the degradation products were transformed into single cell protein (SCP) by the Y. lipolytica cells. After 15 d of fermentation with the whole cell cellulases, the protein content of the maize stover and the rice straw reached 16.23% and 14.75%, which increased by 168.26% and 161.52% compared with the control, respectively. This study provides a new stage for the efficient utilization of stover in the feed industry.

Synergistic effect of cellulase and xylanase during hydrolysis of natural lignocellulosic substrates.

Synergistic combination of cellulase and xylanase has been performed on pre-treated substrates in many previous studies, while few on natural substrates. In this study, three unpretreated lignocellulosic substrates were studied, including corncob, corn stover, and rice straw. The results indicated that when the mixed cellulase and xylanase were applied, reducing sugar concentrations were calculated as 19.53, 15.56, and 17.35mg/ml, respectively, based on the 3,5 dinitrosalicylic acid (DNS) method. Compared to the treatment with only cellulose, the hydrolysis yields caused by mixed cellulase and xylanase were improved by 133%, 164%, and 545%, respectively. In addition, the conversion yield of corncob, corn stover, and rice straw by cellulase-xylanase co-treatment reached 43.9%, 48.5%, and 40.2%, respectively, based on HPLC analysis, which confirmed the synergistic effect of cellulase-xylanase that was much higher than either of the single enzyme treatment. The substrate morphology was also evaluated to explore the synergistic mechanism of cellulase-xylanase.

Simultaneous saccharification and fermentation of corncobs with genetically modified Saccharomyces cerevisiae and characterization of their microstructure during hydrolysis.

Cellulose is an abundant natural polysaccharide that is universally distributed. It can be extracted from corncobs, which are inexpensive, easily accessible, renewable, and environmentally friendly. A common strategy for effectively utilizing cellulose is efficient heterogeneous expression of cellulase genes in Saccharomyces cerevisiae. However, the improvement of cellulose utilization is a relevant issue. Based on our previous findings, we constructed an integrated secretion expression vector, pHBM368-pgk, containing a constitutive promoter sequence. Three genetically modified S. cerevisiae strains containing heterologous ß-glucosidase, exoglucanase, and endoglucanase genes were constructed. The results of a 1-L bioreactor fermentation process revealed that the mixed recombinant S. cerevisiae could efficiently carry out simultaneous saccharification and fermentation (SSF) by using corncobs as the sole carbon source. The ethanol concentration reached 6.37 g/L after 96 hours of fermentation, which was about 3 times higher than that produced by genetically modified S. cerevisiae with the inducible promoter sequence. To investigate the microstructure characteristics of hydrolyzed corncobs during the fermentation process, corncob residues were detected by using a scanning electron microscope. This study provides a feasible method to improve the effect of SSF using corncobs as the sole carbon source.

Systematic screening and characterization of astragalosides in an oral solution of Radix Astragali by liquid chromatography with quadrupole time-of-flight mass spectrometry and Peakview software.

Liquid chromatography with quadrupole time-of-flight mass spectrometry coupled with automated data analysis by Peakview software was employed to systematically screen and characterize the astragalosides in Radix Astragali, a Chinese medical preparation. The separation was performed on a poroshell 120 SB-C18 column equipped in a conventional liquid chromatography system. After being separated using a general gradient elution, the analytes were detected by the triple quadrupole time-of-flight mass spectrometer in both positive- and negative-ion modes. The mass defect filtering function built in the Peakview software was utilized to rapidly screen the potential ions of interest, while some functions of Peakview such as Formula Finder, XIC manager, and IDA Explorer were employed to facilitate the assignment or characterization of the screened astragalosides. A total of 42 astragalosides were screened and tentatively characterized or assigned, and 20 of them were firstly detected in Radix Astragali. According to the screened astragalosides, acetylation, glycosidation, hydrogenation, oxidation, and hydration were considered to be the major secondary metabolic pathways involved in the formation of the astragalosides. The combination of liquid chromatography with quadrupole time-of-flight mass spectrometry and automated Peakview analysis is a feasible and efficient tool to screen and identify the constituents in complex matrices of herbal medicines.

[2-Formyl-4-methyl-6-({2-[2-(4-nitro-benzyl-amino)-ethyl-amino]-ethyl-imino}-meth-yl)phenolato]nickel(II) perchlorate.

In the unsymmetrical title complex, [Ni(C(20)H(23)N(4)O(4))]ClO(4), the coordination geometry for the Ni(II) atom can be described as square planar. The aromatic rings in the two ligands are almost vertical, with a dihedral angle of 85.3°. In the crystal, cations and anions are linked by weak C(N)-H⋯O hydrogen bonding.

Chlorido[N,N'-dibenzyl-N,N'-bis-(pyridin-2-ylmeth-yl)ethane-1,2-diamine]-copper(II) perchlorate methanol monosolvate.

In the title solvated mol-ecular salt, [CuCl(C(28)H(30)N(4))]ClO(4)·CH(3)OH, the Cu(2+) ion is coordinated by the N,N',N'',N'''-tetra-dentate ligand and a chloride ion, generating a very distorted square-based pyramidal CuN(4)Cl coordination geometry with the Cl(-) ion in the basal position. In the crystal, the solvent mol-ecules and anions are linked by weak O-H⋯O hydrogen bonding.

Cell-surface display of the active mannanase in Yarrowia lipolytica with a novel surface-display system.

A novel surface-display system was constructed using the cell-wall anchor protein Flo1p from Saccharomyces cerevisiae, the mannanase (man1) from Bacillus subtilis fused with the C-terminus of Flo1p and the 6xHis tag was inserted between Flo1p and man1. The fusion protein was displayed on the cell surface of Yarrowia lipolytica successfully, and it was confirmed by immunofluorescence. In succession, the surface-displayed mannanase was characterized. The optimum catalytic conditions for the recombinant mannanase were 55 degrees C at pH 6.0, and it exhibited high stability against pH variation. The highest activity of the recombinant mannanase reached 62.3 IU/g (dry cell weight) after the recombinant was cultivated for 96 h in YPD medium [1% (w/v) yeast extract/2% (w/v) peptone/2% (w/v) glucose]. To our knowledge, the present paper is the first to report that high-activity mannanase is displayed on the cell surface of Y. lipolytica with Flo1p.

Cell surface display of functionally active lipases from Yarrowia lipolytica in Pichia pastoris.

The lipase genes of Yarrowia lipolytica, LIPY7 and LIPY8, fused with FLO-flocculation domain sequence from Saccharomyces cerevisiae at their N-termini, were expressed in Pichia pastoris KM71. Following the induction with methanol, the recombinant proteins were displayed on the cell surface of P. pastoris, as confirmed by the confocal laser scanning microscopy. The LipY7p and LipY8p were anchored on P. pastoris via the flocculation functional domain of Flo1p. The surface-displayed lipases were characterized for their application as the whole-cell biocatalyst. These lipases can also be cleaved off from their anchor by enterokinase treatment to yield functionally active proteins in the supernatant offering an alternative purification method for LipY7p and LipY8p.

Expression and purification of two lipases from Yarrowia lipolytica AS 2.1216.

We isolated two lipase genes LIPY7, LIPY8 from Yarrowia lipolytica CGMCC (China general microbiological culture collection center) AS 2.1216. The LIPY7 and LIPY8 genes encode a 366 and a 371-amino acid protein, respectively. The lipase genes with 6 x His tag sequence were cloned into expression vector pPIC9K and successfully integrated into a heterologous fungal host Pichia pastoris KM71, respectively. The recombinants were induced by methanol to secrete active lipases into cultural medium. The recombinant lipases were also purified and characterized.

[Secreted expression of Bacillus pumilus xylanase gene in Pichia pastoris and study on enzymatic properties].

The endo-1,4-xylanase gene from Bacillus pumilus HB030 was cloned into the Pichia pastoris expression vector, pPIC9k, the recombinant plasmid was named pHBM220. The digested recombinant plasmid pHBM220 was transformed into Pichia pastoris KM71, GS115, SMD1168, respectively. The recombinant Pichia pastoris KM71 (pHBM220), GS115 (pHBM220), SMD1168 (pHBM220) secreted functional endo-1,4-xylanase, and the enzymatic activities reached 10.80IU/mL, 11.63IU/mL, 9.68IU/mL, respectively. The temperature and pH optimum for the recombinant xylanase were 60 degrees C and pH5.5, respectively.