Symmetric and Asymmetric Zeolitic Imidazolate Frameworks (ZIFs)/Polybenzimidazole (PBI) Nanocomposite Membranes for Hydrogen Purification at High Temperatures

High‐performance zeolitic imidazolate frameworks (ZIFs)/polybenzimidazole (PBI) nanocomposites are molecularly designed for hydrogen separation at high temperatures, and demonstrate it in a useful configuration as dual‐layer hollow fibers for the first time. By incorporating as‐synthesized nanoporous ZIF‐8 nanoparticles into the high thermal stability but extremely low permeability polybenzimidazole (PBI), the resultant mixed matrix membranes show an impressive enhancement in H₂ permeability as high as a hundred times without any significant deduction in H₂/CO₂ selectivity. The 30/70 ZIF‐8/PBI dense membrane has a H₂ permeability of 105.4 Barrer and a H₂/CO₂ selectivity of 12.3. This performance is far superior to ZIF‐7/PBI membranes and is the best ever reported data for H₂‐selective polymeric materials in the literature. Meanwhile, defect‐free ZIF‐8‐PBI/Matrimid dual‐layer hollow fibers are successfully fabricated, without post‐annealing and coating, by optimizing ZIF‐8 nanoparticle loadings, spinning conditions, and solvent‐exchange procedures. Two types of hollow fibers targeted at either high H₂/CO₂ selectivity or high H₂ permeance are developed: i) PZM10‐I B fibers with a medium H₂ permeance of 64.5 GPU (2.16 ×10^?8 mol m^?2 s^?1 Pa^?1) at 180°C and a high H₂/CO₂ selectivity of 12.3, and, ii) PZM33‐I B fibers with a high H₂ permeance of 202 GPU (6.77 ×10^?8 mol m^?2 s^?1 Pa^?1) at 180°C and a medium H₂/CO₂ selectivity of 7.7. This work not only molecularly designs novel nanocomposite materials for harsh industrial applications, such as syngas and hydrogen production, but also, for the first time, synergistically combines the strengths of both ZIF‐8 and PBI for energy‐related applications.     

Bimetallic Cobalt‐Based Phosphide Zeolitic Imidazolate Framework: CoPx Phase‐Dependent Electrical Conductivity and Hydrogen Atom Adsorption Energy for Efficient Overall Water Splitting

Cobalt‐based bimetallic phosphide encapsulated in carbonized zeolitic imadazolate frameworks has been successfully synthesized and showed excellent activities toward both hydrogen evolution reaction (HER) and oxygen evolution reaction (OER). Density functional theory calculation and electrochemical measurements reveal that the electrical conductivity and electrochemical activity are closely associated with the Co₂P/CoP mixed phase behaviors upon Cu metal doping. This relationship is found to be the decisive factor for enhanced electrocatalytic performance. Moreover, the precise control of Cu content in Co‐host lattice effectively alters the Gibbs free energy for H* adsorption, which is favorable for facilitating reaction kinetics. Impressively, an optimized performance has been achieved with mild Cu doping in Cu_0.3Co_2.7P/nitrogen‐doped carbon (NC) which exhibits an ultralow overpotential of 0.19 V at 10 mA cm^–2 and satisfying stability for OER. Cu_0.3Co_2.7P/NC also shows excellent HER activity, affording a current density of 10 mA cm^–2 at a low overpotential of 0.22 V. In addition, a homemade electrolyzer with Cu_0.3Co_2.7P/NC paired electrodes shows 60% larger current density than Pt/RuO₂ couple at 1.74 V, along with negligible catalytic deactivation after 50 h operation. The manipulation of electronic structure by controlled incorporation of second metal sheds light on understanding and synthesizing bimetallic transition metal phosphides for electrolysis‐based energy conversion.     

Dye-ligand membranes as selective adsorbents for rapid purification of enzymes: a case study

A new adsorbent for the selective binding of enzymes, in the form of microporous membranes carrying triazine dyes as pseudo-affinity ligand, has been implemented in the recovery of glucose-6-phosphate dehydrogenase from yeast. A detailed investigation of the process parameters has been performed. In the adsorption step, the contact time for binding G6PDH could be reduced down to 0.25 s without significant decrease of the capture efficiency. Hence, fast filtration allowed to isolate G6PDH from a dilute extract (1.6 mug G6PDH . mL(-1)), where the enzyme accounted for 1% of the proteins. The yield of the selective elution step using NADP was only 70% at best. It could be improved to near 100% by supplementing the eluent with ethylene glycol, without loss of selectivity. A Scale-up of the cross-section of the membrane by a factor of 40 allowed to purify 1140 U from 0.6 L extract from 1% to 57% purity with 82% yield, within 10 minutes. The case study presented here demonstrates the applicability of general-purpose membrane adsorbents for the purification of enzymes.     

Development of an expanded bed technique for an affinity purification of G6PDH from unclarified yeast cell homogenates

The use of an expanded bed of STREAMLINE Red H-7B for the purification of the intracellular glycolytic enzyme glucose 6-phosphate dehydrogenase (G6PDH) directly from untreated preparations of disrupted yeast cells has been investigated. Small-scale experiments, carried out in packed beds, have shown that the optimal pH for adsorption is 6.0 and have enabled optimization of elution conditions using a series of eluents. The dynamic capacity of the adsorbent for G6PDH was determined in a small expanded bed to be 28 units/mL. These results were used to develop a preparative scale separation of G6PDH in a STREAMLINE 50 expanded bed column. G6PDH was purified directly from an unclarified yeast homogenate in 99% yield with an average purification factor in the eluted fraction of 103. Cleaning-in-place (CIP) procedures using 0.5 M NaOH and 4M urea in 60% (v/v) ethanol have demonstrated that the adsorbent can be regenerated with no loss of adsorption capacity of alteration of bed expansion characteristics after many cycles of operation. (c) 1995 John Wiley & Sons, Inc.     

聚合高分子电解质分离纯化蛋白质

聚合高分子电解质含有大量阳离子或阴离子,通过静电作用结合带相反电荷的蛋白质,生成聚合高分子电解质-蛋白质复合物,电解质-蛋白质复合物通过桥连作用或疏水作用形成沉淀颗粒;聚合高分子电解质的选择性沉淀作用受电解质的分子量、添加剂量、溶液离子强度和pH的影响。用高分子电解质从大规模的低浓度溶液中选择性地沉淀目的蛋白质,为生物工程的下游处理开辟了一条新途径。     

Calorimetric and molecular simulation study on unfrozen water characteristics in aqueous sugar solutions: implications for biopreservation

Intensive studies have been conducted to determine the protective mechanisms of sugars that have proven beneficial to the biopreservation application. However, little has been known about the unfrozen water content that aqueous sugar solutions can possess when frozen at cryogenic temperatures. This study conducted calorimetric measurements to determine the unfrozen water content in frozen aqueous solutions of glucose, fructose, sucrose and trehalose of multiple concentrations. The hydrogen-bonding network in these solutions was characterised by molecular simulations. The experimental results showed that more water could be prevented from ice crystallisation in a more concentrated solution. Disaccharides, especially trehalose, are more effective than other protectants (e.g., glucose, glycerol and dimethyl sulfoxide) for detaining water in the unfrozen state. Moreover, it was found that, at molecular levels, there were more hydrogen bonds between sugar and water molecules in a more concentrated solution. From both macro- and microscopic perspectives, trehalose was demonstrated to be a much more effective cryoprotectant than others. This comparative study proved that the unfrozen water should be mainly attributed to hydrogen bonds between sugar and water in the mixture. Our findings will provide valuable information for determining the physical state of cryopreserved biomatrix and guiding the preparation of protective formulations.     

Highly diverse variable number tandem repeat loci in the E. coli O157:H7 and O55:H7 genomes for high-resolution molecular typing

AIM: Evaluation of the Escherichia coli genome for variable number tandem repeat (VNTR) loci in order to provide a subtyping tool with greater discrimination and more efficient capacity. METHODS AND RESULTS: Twenty-nine putative VNTR loci were identified from the E. coli genomic sequence. Their variability was validated by characterizing the number of repeats at each locus in a set of 56 E. coli O157:H7/HN and O55:H7 isolates. An optimized multiplex assay system was developed to facility high capacity analysis. Locus diversity values ranged from 0.23 to 0.95 while the number of alleles ranged from two to 29. This multiple-locus VNTR analysis (MLVA) data was used to describe genetic relationships among these isolates and was compared with PFGE (pulse field gel electrophoresis) data from a subset of the same strains. Genetic similarity values were highly correlated between the two approaches, through MLVA was capable of discrimination amongst closely related isolates when PFGE similar values were equal to 1.0. CONCLUSIONS: Highly variable VNTR loci exist in the E. coli O157:H7 genome and are excellent estimators of genetic relationships, in particular for closely related isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: Escherichia coli O157:H7 MLVA offers a complimentary analysis to the more traditional PFGE approach. Application of MLVA to an outbreak cluster could generate superior molecular epidemiology and result in a more effective public health response.     

Characterization of inducible stx2-positive Escherichia coli O157:H7/H7- strains isolated from cattle in France

Aims:  To quantify the variability of the Shiga toxin 2 (Stx2) production by a panel of stx2 -positive Escherichia coli O157:H7/H7- isolates from healthy cattle before and after induction with enrofloxacin.
Methods and Results:  ProSpecT^® ELISA was used to quantify the Stx2 production by stx2 -positive E. coli O157:H7/H7- isolates in native conditions (basal level) or after induction with enrofloxacin. Whereas only 15·2% of the E. coli O157:H7/H7- strains studied displayed significant amounts of detectable Stx2 without induction, most of them were shown to be inducible, and at various levels, in presence of subinhibitory concentrations of enrofloxacin.
Conclusions:  We demonstrated the capability of a highly elevated proportion of stx2 -positive, but constitutively Stx2 -negative, E. coli O157:H7/H7- isolates from healthy cattle to produce significant levels of Shiga toxin Stx2 in presence of subtherapeutic concentrations of enrofloxacin, an antibiotic of the fluoroquinolones family only licensed for veterinary use.
Significance and Impact of the Study:  This study documents the risk that bovine-associated Shiga toxin producing E. coli isolates may become more frequently pathogenic to humans as a side-effect of the increasing use of veterinary fluoroquinolones in the oral treatment of food animals like cattle or poultry.     

Dose-Response Envelope for Escherichia coli O157:H7

Escherichia coli O157:H7 is an emerging food and waterborne pathogen in the U.S. and internationally. The objective of this work was to develop a dose-response model for illness by this organism that bounds the uncertainty in the dose-response relationship. No human clinical trial data are available for E. coli O157:H7, but such data are available for two surrogate pathogens: enteropathogenic E. coli (EPEC) and Shigella dysenteriae. E. coli O157:H7 outbreak data provide an initial estimate of the most likely value of the dose-response relationship within the bounds of an envelope defined by beta-Poisson dose-response models fit to the EPEC and S. dysenteriae data. The most likely value of the median effective dose for E. coli O157:H7 is estimated to be approximately 190[emsp4 ]000 colony forming units (cfu). At a dose level of 100[emsp4 ]cfu, the median response predicted by the model is six percent.     

An attempt to design an isoluminol-hydrogen peroxidase-amplified CL that measures intracellularly produced H2O2 in phagocytes: sensitivity for H2O2 is not high enough to allow detection.

A technique was designed for the determination of hydrogen peroxide release. We found, however, that the isoluminol-amplified chemiluminescence technique is a suitable tool for measuring secretion of superoxide but not hydrogen peroxide.     

Molecular simulation study of the quantum effects of hydrogen adsorption in metal-organic frameworks: influences of pore size and temperature

In this work a systematic molecular simulation study was performed to investigate the influence of pore size and temperature on the quantum effects of hydrogen adsorption in metal-organic frameworks (MOFs) with temperature varied from 40 to 120 K. To do this, three isoreticular MOFs (IRMOFs) with different pore sizes were adopted, and quadratic Feynman–Hibbs (FH) effective potential was introduced to consider the quantum effect. The results show that quantum effects diminish with increasing pore size of IRMOFs at lower pressure (loading), while the opposite trend appears at higher pressure (loading). Through the simulations it is also found that the quantum effects may be dominantly determined by the adsorbate–adsorbate or adsorbate–MOFs interactions with the varying of pressure (loading). In addition, the results also indicate how the temperature influences the quantum effects of H₂ adsorption in MOFs within the pressure range considered.     

Precursors for Cholesterol Synthesis (7-Dehydrocholesterol, 7-Dehydrodesmosterol, and Desmosterol): Cholesterol/7-Dehydrocholesterol Ratio as an Index of Development and Aging in PNS but Not in CNS

In rat sciatic nerve, the 7-dehydrocholesterol content decreased dramatically during the postnatal period and slowly during adulthood and aging. In contrast, the 7-dehydrodesmosterol content peaked at 14 days and was nearly undetectable after 60 days. The desmosterol content peaked at 21 days and was nearly undetectable after 1 year. The cholesterol content increased up to 21 days and remained nearly constant thereafter. In brain (in contrast to sciatic nerve), 7-dehydrodesmosterol and desmosterol contents decreased dramatically during development and slightly during adulthood and aging; the 7-dehydrocholesterol content peaked at 21 days and remained constant during aging. Only 7-dehydrocholesterol was dramatically more concentrated in PNS than in CNS. In brain, the cholesterol/7-dehydrocholesterol ratio increased during development and remained stable after 6 months. In contrast, in sciatic nerve, this ratio continuously increased during development and aging (950-fold between 5 days and 18 months). Thus, the cholesterol/7-dehydrocholesterol ratio is a useful biochemical index of development and aging in the PNS.     

Novel N-biphenyl-2-ylmethyl 2-methoxyphenylpiperazinylalkanamides as 5-HT7R antagonists for the treatment of depression

5-HT₇ receptor (5-HT₇R) is a promising target for the treatment of depression and neuropathic pain. 5-HT₇R antagonists exhibited antidepressant effects, while the agonists produced strong anti-hyperalgesic effects. In our efforts to discover selective 5-HT₇R antagonists or agonists, N-biphenylylmethyl 2-methoxyphenylpiperazinylalkanamides 1 were designed, synthesized, and biologically evaluated against 5-HT₇R. Among the synthesized compounds, N-2′-chlorobiphenylylmethyl 2-methoxyphenylpiperazinylpentanamide 1–8 showed the best binding affinity with a K_i value of 8.69 nM and it was verified as a novel antagonist according to functional assays. The compound 1–8 was very selective over 5-HT_1DR, 5-HT_2AR, 5-HT₃R, 5-HT_5AR and 5-HT₆R and moderately selective over 5-HT_1AR, 5-HT_1BR and 5-HT_2CR. The novel 5-HT₇R antagonist 1–8 exhibited an antidepressant effect at a dose of 25 mg/kg in the forced swimming test in mice and showed a U-shaped dose–response curve which typically appears in 5-HT₇R antagonists such as SB-269970 and lurasidone.     

Exploring purine N7 interactions via atomic mutagenesis: the group I ribozyme as a case study

Atomic mutagenesis has emerged as a powerful tool to unravel specific interactions in complex RNA molecules. An early extensive study of analogs of the exogenous guanosine nucleophile in group I intron self-splicing by Bass and Cech demonstrated structure-function relationships analogous to those seen for protein ligands and provided strong evidence for a well-formed substrate binding site made of RNA. Subsequent functional and structural studies have confirmed these interacting sites and extended our understanding of them, with one notable exception. Whereas 7-methyl guanosine did not affect reactivity in the original study, a subsequent study revealed a deleterious effect of the seemingly more conservative 7-deaza substitution. Here we investigate this paradox, studying these and other analogs with the more thoroughly characterized ribozyme derived from the Tetrahymena group I intron. We found that the 7-deaza substitution lowers binding by ~20-fold, relative to the cognate exogenous guanosine nucleophile, whereas binding and reaction with 7-methyl and 8-aza-7-deaza substitutions have no effect. These and additional results suggest that there is no functionally important contact between the N7 atom of the exogenous guanosine and the ribozyme. Rather, they are consistent with indirect effects introduced by the N7 substitution on stacking interactions and/or solvation that are important for binding. The set of analogs used herein should be valuable in deciphering nucleic acid interactions and how they change through reaction cycles for other RNAs and RNA/protein complexes.     

Response-surface methodology for the production and the purification of a new H2O2-tolerant alkaline protease from Bacillus invictae AH1 strain

This work deals with the optimization of the culture conditions of Bacillus invictae AH1 in order to increase the production level of the proteolytic activity. Response-surface methodology (RSM) was applied for the most significant fermentation parameters (concentration of wheat bran and K₂HPO₄/KH₂PO₄) that were earlier identified by Plackett–Burman Design from seven possible factors. A central composite design was used and the quadratic regression model of producing active protease was built. A maximum protease activity was reached and validated experimentally, using a maximum wheat bran concentration (50 g/L) with increased K₂HPO₄/KH₂PO₄ concentration (2.275 g/L). Protease production obtained experimentally coincident with the predicted value and the model was proven to be adequate. Interestingly, the use of RSM increased the protease production by four times (7,000 U/mL) using a low-cost substrate and a culture time of 40 hr, as compared to the standard culture conditions. In the second part of this study, a H₂O₂-tolerant alkaline protease produced from B. invictae AH1 with a molecular mass of about 41 kDa, noted P3, was purified by successive steps of ultrafiltration, gel filtration and ion exchange chromatography. The K _m and V_max values of the purified protease using casein, as substrate, were about 4 mg/mL and 27 μM/min, respectively. The highest enzyme activity was found at pH 9.0 and a temperature of 60°C. In addition, the enzyme showed a quasi-total stability against H₂O₂ (5% for 1 hr) and against most of the tested solid and liquid detergents, suggesting its eventual use in bio-detergent formulations.     

Comparison of an immunochromatographic method and the TaqMan E. coli O157:H7 assay for detection of Escherichia coli O157:H7 in alfalfa sprout spent irrigation water and in sprouts after blanching

An immunochromatographic-based assay (Quixtrade mark E. coli O157 Sprout Assay) and a polymerase chain reaction (PCR)-based assay (TaqMan E. coli O157:H7 Kit) were used to detect Escherichia coli O157:H7 strain 380-94 in spent irrigation water from alfalfa sprouts grown from artificially contaminated seeds. Ten, 25, 60, or 100 seeds contaminated by immersion for 15 min in a suspension of E. coli O157:H7 at concentrations of 10(6) or 10(8) cfu/ml were mixed with 20 g of non-inoculated seeds in plastic trays for sprouting. The seeds were sprayed with tap water for 15 s every hour and spent irrigation water was collected at intervals and tested. E. coli O157:H7 was detected in non-enriched water by both the TaqMan PCR (30 of 30 samples) and the immunoassay (9 of 24 samples) in water collected 30 h from the start of the sprouting process. However, enrichment of the spent irrigation water in brain heart infusion (BHI) broth at 37 degrees C for 20 h permitted detection of E. coli O157:H7 in water collected 8 h from the start of sprouting using both methods, even in trays containing as few as 10 inoculated seeds. The TaqMan PCR assay was more sensitive (more positive samples were observed earlier in the sprouting process) than the immunoassay; however, the immunoassay was easier to perform and was more rapid. At 72 h after the start of the sprouting process, the sprouts were heated at 100 degrees C for 30 s to determine the effectiveness of blanching for inactivation of E. coli O157:H7. All of the 32 samples tested with the TaqMan assay and 16 of 32 samples tested with the Quixtrade mark assay gave positive results for E. coli O157:H7 after enrichment of the blanched sprouts at 37 degrees C for 24 h. In addition, the organism was detected on Rainbow Agar O157 in 9 of 32 samples after 24 h of enrichment of the blanched sprouts. In conclusion, E. coli O157:H7 was detected in spent irrigation water collected from sprouts grown from artificially contaminated seeds by both the TaqMan and Quixtrade mark assays. The data also revealed that blanching may not be effective to completely inactivate all the E. coli O157:H7 that may be present in sprouts.     

Evaluation of AFLP, a high-resolution DNA fingerprinting method, as a tool for molecular subtyping of enterohemorrhagic Escherichia coli O157:H7 isolates.

The amplified fragment-length polymorphism (AFLPTM) technique is based on the selective PCR amplification of restriction fragments. We investigated the utility of AFLP in the molecular subtyping of enterohemorrhagic Escherichia coli serotype O157:H7 isolates. We analyzed a total of 46 isolates of E. coli O157:H7 along with other serotypes, O26:H11, 0114:H19 and 0119:NT. Isolates of E. coli O157:H7 derived from the same outbreak showed an identical AFLP-banding pattern and were subtyped into the same group, giving results almost consistent with those of a pulsed-field gel electrophoresis (PFGE) study, while other serotypes showed clearly different patterns from those of E. coli O157:H7. These results suggest that the AFLP technique has potential as an alternative tool for the molecular epidemiology of E. coli O157:H7.