Engineered Amp C β-lactamase as a fluorescent screening tool for class C β-lactamase inhibitors

Class C β-lactamases mediate antibiotic resistance in bacteria by efficiently hydrolyzing a broad range of β-lactam antibiotics. With their clinical significance and the lack of commercially available effective inhibitors, development of class C β-lactamase inhibitors has become one of the recent hot issues in the pharmaceutical industry. In this paper, we report the protein engineering of a fluorescent Amp C β-lactamase mutant designated as V211Cf for the in vitro screening of class C β-lactamase inhibitors. When a fluorescein (f) was incorporated at the entrance of the enzyme's active site (position 211), Amp C β-lactamase from Enterobacter cloacae P99 was tailor-made into a novel fluorescent biosensing protein that could display a fluorescence enhancement upon binding with its β-lactam substrates/inhibitors. With its catalytic activity close to the wild-type level, V211Cf can act as a "natural" fluorescent drug target for screening small binding molecules. In addition, V211Cf can allow specific detection for its active-site binding molecules and discriminate them from nondruglike molecules in the screen. Furthermore, V211Cf is amenable to a high throughput format. Taken together, V211Cf demonstrates the potential as an efficient tool for screening class C β-lactamase inhibitors and facilitates the discovery of therapeutics that can combat the clinically important class C β-lactamases.     

Heterobimetallic Ru(II)-Eu(III) complex as chemodosimeter for selective biogenic amine odorants detection in fish sample

Gaseous biogenic amines such as putrescine, spermidine, aniline, and trimethylamine are important biomolecules that play many crucial roles in metabolism and medical diagnostics. A chemodosimetric detection assay has been developed for those gaseous amines by Ru(II)-Eu(III) heterobimetallic complexes, K{[Ru(II)((t)Bubpy)(CN)(4)](2)Eu(III)(H(2)O)(4)} (where (t)Bubpy = 4,4'-di-tert-butyl-2,2'-bipyridine). Synthesis, X-ray crystal characterization, and spectroscopic properties of this Ru(II)-Eu(III) heterobimetallic complex were reported. Binding properties of the Ru(II)-Eu(III) complex with common gases revealed that this complex is very selective to gaseous amine molecules. Sensitivity of this complex toward the amines was found as ～log k() = 4.5-4.8. Real time monitoring of gaseous biogenic amines was applied to real fish samples (Atlantic mackerel) by studying the spectrofluorimetric responses of the Ru(II)-Eu(III) complex toward different biogenic amine concentration. GC/MS studies were also used as a reference for the studies. A linear spectrofluorimetric response was found toward biogenic amine concentration in real fish samples. This complex was found to respond specifically to those biogenic amines down to 10 ppb.     

HILIC-UPLC-MS for exploratory urinary metabolic profiling in toxicological studies

Hydrophilic interaction ultra performance liquid chromatography (HILIC-UPLC) permits the analysis of highly polar metabolites, providing complementary information to reversed-phase (RP) chromatography. HILIC-UPLC-TOF-MS was investigated for the global metabolic profiling of rat urine samples generated in an experimental hepatotoxicity study of galactosamine (galN) and the concomitant investigation of the protective effect of glycine. Within-run repeatability and stability over a large sample batch (>200 samples, 60 h run-time) was assessed through the repeat analysis of a quality control sample. Following system equilibration, excellent repeatability was observed in terms of retention time (CV < 1.7%), signal intensity (CV < 14%), and mass variability (<0.005 amu), providing a good measure of reproducibility. Classification of urinary metabolic profiles according to treatment was observed, with significant changes in specific metabolites after galN exposure, including increased urocanic acid, N-acetylglucosamine, and decreased 2-oxoglutarate. A novel finding from this HILIC-UPLC-MS approach was elevated urinary tyramine in galN-treated rats, reflecting disturbed amino acid metabolism. These results show HILIC-UPLC-MS to be a promising method for global metabolic profiling, demonstrating high within-run repeatability, even over an extended run time. Retention of polar endogenous analytes and xenobiotic metabolites was improved compared with RP studies, including galN, N-acetylglucosamine, oxoglutarate, and urocanic acid, enhancing metabolome coverage and potentially improving biomarker discovery.     

Analysis for a two-dissimilar-component cold standby repairable system with repair priority

In this paper, a cold standby repairable system consisting of two dissimilar components and one repairman is studied. Assume that working time distributions and repair time distributions of the two components are both exponential, and Component 1 has repair priority when both components are broken down. After repair, Component 1 follows a geometric process repair while Component 2 obeys a perfect repair. Under these assumptions, using the perfect repair model, the geometric process repair model and the supplementary variable technique, we not only study some important reliability indices, but also consider a replacement policy T, under which the system is replaced when the working age of Component 1 reaches T. Our problem is to determine an optimal policy T^? such that the long-run average loss per unit time (i.e. average loss rate) of the system is minimized. The explicit expression for the average loss rate of the system is derived, and the corresponding optimal replacement policy T^? can be found numerically. Finally, a numerical example for replacement policy T is given to illustrate some theoretical results and the model's applicability.     

Large strain response in acceptor- and donor-doped Bi<Subscript>0.5</Subscript>Na<Subscript>0.5</Subscript>TiO<Subscript>3</Subscript>-based lead-free ceramics

Effects of Fe and La addition on the dielectric, ferroelectric, and piezoelectric properties of Bi_0.5Na_0.5TiO₃–Bi_0.5Li_0.5TiO₃–BaTiO₃–Mn ceramics were investigated. Similar to the doping effect in lead-based piezoelectric materials, here the Fe-doped ceramic created a hard effect with an improved mechanical quality factor (Q _m) ~ 160, coercive field (E _c) ~ 2.9 kV/mm, decreased dielectric constant ( e₃₃^T /e₀ ) ~ 80 3, \left( {\varepsilon_{33}^{T} /\varepsilon_{0} } \right)\sim 80 3, and loss (tanδ) ~ 0.024 while the La-doped one indicated a soft feature with improved piezoelectric constant (d ₃₃) ~ 184 pC/N, e₃₃^T /e₀   ~ 983, \varepsilon_{33}^{T} /\varepsilon_{0} \,\sim { 983}, tanδ ~ 0.033, and decreased E _c ~ 2.46 kV/mm. In addition, the temperature dependence of the ferroelectric hysteresis loops and strain response under unipolar electric field was also studied. Around the depolarization temperature T _d, large strain value was obtained with the normalized d₃₃^* d_{33}^{*} up to ~1,000 pC/N, which was suggested originated from the development of the short-range order or non-polar phases in the ferroelectric matrix. All these would provide a new way to realize high piezoelectric response for practical application in different temperature scale.     

Flow of gas—solid mixtures in standpipes. A review

In this paper, the literature on standpipe flow is systematically reviewed. In particular, four possible flow regimes are defined, viz. dense phase fluidized solids flow (DENFLO), lean phase fluidized solids flow (LEANFLO), transition packed bed flow (TRANPACFLO) and packed bed flow (PACFLO). The possibilities of coexistence of more than one of these flow regimes and the prediction of these coexistences are discussed. Available equations pertaining to each flow regime and to flow of gas—solids mixture through a slide valve are summarized. The use of the equations for analysis of industrial standpipe operation is illustrated by means of two case studies. Finally, areas where further work would be fruitful are pinpointed. More experimental work is urged to verify the validity of some of the equations presented and to improve our understanding of the different types of flow instabilities in standpipe operation.     

An analysis of moving-bed flow of solids down standpipes and side valves

A method of analysing solids downflow through standpipe and slide valve in the moving bed mode is presented. The method is based on combining Yoon and Kunii equations for moving-bed flow with a proposed modified orifice equation for gas—solid flow. The analysis correctly predicts the observed trend of the effects of gas injection into the standpipe. It also provides quantitative prediction of the conditions for transition from moving-bed flow to dense phase fluidized solids flow and to lean phase “streaming flow”. The latter flow pattern refers to solids streaming down a tube at high voidage typified by the discharge of solid down a tube with no constriction at the lower end of the tube. Further work is required to test the general applicability of the present analysis and to extend the analysis to yield a generalized quantitative flow regime diagram for downflow of solids in a pipe.