KMgF3:Ni2+晶体的占位、局部结构、吸收光谱和顺磁g因子的统一解释

利用完全对角化方法和强场耦合方案,采用半自洽场(semi-SCF)自由Ni2+的d轨道模型和Ni2+-6X-(x=F,Cl,Br,I)络合物的μ-κ-α模型研究,建立了含有过渡族金属离子的晶体的局域结构与吸收光谱和顺磁g因子之间的定量关系,对KMgF3:Ni2+晶体的占位、局域结构、吸收光谱和顺磁g因子作出了统一解释,预测了KMgF3:Ni2+晶体的光谱精细结构.所得理论计算结果与实验值符合得很好.     

Adsorption of detergent-solubilized and phospholipase C-solubilized alkaline phosphatase at air/liquid interfaces

To investigate the influence of a hydrophobic anchor on protein adsorption, equilibrium and dynamic aspects of the adsorption of two different solubilized forms of rat osseous plate alkaline phosphatase on Langmuir monolayers of dimyristoylphosphatidic acid (DMPA) were studied. Surface pressure and surface potential measurements at air/liquid interfaces were carried out using the detergent-solubilized form (DSAP) of alkaline phosphatase, which holds a glycosylphosphatidylinositol (GPI) hydrophobic anchor, and the glycosylphosphatidylinositol-specific phospholipase C-solubilized form (PLSAP), lacking the GPI anchor. Similar surface transitions observed for both DMPA and DMPA/PLSAP mixed monolayers indicate that the presence of PLSAP does not promote significant changes in surface packing of the DMPA monolayer. However, PLSAP interacts with the polar portion of the phospholipid even at high lateral compression. The presence of the GPI anchor increases the adsorption of DSAP at a plain air/liquid interface and also enables the penetration of the protein into the DMPA monolayers. The penetration is dependent on both time and surface pressure. Up to 20 mN/m, the surface pressure increases smoothly indicating a diffusion followed by an adsorption process. Above 20 mN/m, after a fast increase, the surface pressure slowly decays to equilibrium values quite close to the initial surface pressures. The results indicate that the molecular packing of the lipid layer drives the enzyme adsorption to the interface either through the GPI anchor or by the polypeptide moiety.     

Olefin epoxidation with tert-butyl hydroperoxide catalyzed by MoO2X2L complexes: a DFT mechanistic study

DFT calculations suggest that the catalytic epoxidation of olefins by Mo(vi) complexes, modeled by MoO2Br2(MeN=C(H)-C(H)=NMe), in the presence of MeOOH, the model for tert-butyl hydroperoxide, starts with a hydrogen transfer from the peroxide to one of the terminal Mo=O oxygen atoms and the remaining MeOO anion binds as a seventh ligand, forming a five-membered Mo-O(alpha)-O(beta)(Me)...H-O-Mo ring held together by a hydrogen bond. In the second step, a concerted approach of ethylene to the Mo-O(alpha) bond gives rise to an intermediate containing a seven-membered Mo-C-C-O(alpha)-O(beta)(Me)...H-O-Mo ring. In the final step, decomposition of the intermediate leads to the starting complex, alcohol and the epoxide. The activation energy for the addition of the olefin (second step) is the highest one, in agreement with available kinetic studies showing that the catalyst formation is not always a rate-limiting step. DFT calculations also show that the alcohol by-product (MeOH) can react with the starting complex, competing with ROOH and hence leading to the progressive catalyst poisoning, which has been observed experimentally.     

Microspot-based ELISA in microfluidics: chemiluminescence and colorimetry detection using integrated thin-film hydrogenated amorphous silicon photodiodes

Microfluidic technology has the potential to decrease the time of analysis and the quantity of sample and reactants required in immunoassays, together with the potential of achieving high sensitivity, multiplexing, and portability. A lab-on-a-chip system was developed and optimized using optical and fluorescence microscopy. Primary antibodies are adsorbed onto the walls of a PDMS-based microchannel via microspotting. This probe antibody is then recognised using secondary FITC or HRP labelled antibodies responsible for providing fluorescence or chemiluminescent and colorimetric signals, respectively. The system incorporated a micron-sized thin-film hydrogenated amorphous silicon photodiode microfabricated on a glass substrate. The primary antibody spots in the PDMS-based microfluidic were precisely aligned with the photodiodes for the direct detection of the antibody-antigen molecular recognition reactions using chemiluminescence and colorimetry. The immunoassay takes ~30 min from assay to the integrated detection. The conditions for probe antibody microspotting and for the flow-through ELISA analysis in the microfluidic format with integrated detection were defined using antibody solutions with concentrations in the nM-μM range. Sequential colorimetric or chemiluminescence detection of specific antibody-antigen molecular recognition was quantitatively detected using the photodiode. Primary antibody surface densities down to 0.182 pmol cm(-2) were detected. Multiplex detection using different microspotted primary antibodies was demonstrated.     

Binding and elution strategy for improved performance of arginine affinity chromatography in supercoiled plasmid DNA purification

New interesting strategies for plasmid DNA (pDNA) purification were designed, exploiting affinity interactions between amino acids and nucleic acids. The potential application of arginine-based chromatography to purify pDNA has been recently described in our work; however, to achieve higher efficiency and selectivity in arginine affinity chromatography, it is essential to characterize the behaviour of binding/elution of supercoiled (sc) isoforms. In this study, two different strategies based on increased sodium chloride (225-250 mm) or arginine (20-70 mm) stepwise gradients are described to purify sc isoforms. Thus, it was proved that well-defined binding/elution conditions are crucial to enhance the purification performance, resulting in an improvement of the final plasmids yields and transfection efficiency, as this could represent a significant impact on therapeutic applications of the purified sc isoform.     

Antiproliferative Effect of Colonic Fermented Phenolic Compounds from Jaboticaba (Myrciaria trunciflora) Fruit Peel in a 3D Cell Model of Colorectal Cancer

Jaboticaba is a Brazilian native berry described as a rich source of phenolic compounds (PC) with health promoting effects. PC from jaboticaba peel powder (JPP) have low intestinal bio-accessibility and are catabolized by gut microbiota. However, the biological implication of PC-derived metabolites produced during JPP digestion remains unclear. This study aimed to evaluate the antiproliferative effects of colonic fermented JPP (FJPP) in a 3D model of colorectal cancer (CRC) composed by HT29 spheroids. JPP samples fermented with human feces during 0, 2, 8, 24 or 48 h were incubated (10,000 µg mL⁻¹) with spheroids, and cell viability was assessed after 72 h. Chemometric analyses (cluster and principal component analyses) were used to identify the main compounds responsible for the bioactive effect. The antiproliferative effect of FJPP in the CRC 3D model was increased between 8 h and 24 h of incubation, and this effect was associated with HHDP-digalloylglucose isomer and dihydroxyphenyl-γ-valerolactone. At 48 h of fermentation, the antiproliferative effect of FJPP was negligible, indicating that the presence of urolithins did not improve the bioactivity of JPP. These findings provide relevant knowledge on the role of colonic microbiota fermentation to generate active phenolic metabolites from JPP with positive impact on CRC.     

Surface density as a significant parameter for the enzymatic activity of two forms of alkaline phosphatase immobilized on phospholipid Langmuir-Blodgett films

Rat osseous plate alkaline phosphatase, a glycosylphosphatidylinositol (GPI)-anchored phosphomonohydrolase, was immobilized on Langmuir-Blodgett (LB) films. Enzyme solubilization either with polyoxyethylene-9-lauryl ether or with a glycosylphosphatidylinositol-specific phospholipase C resulted in a GPI-anchor-containing and a GPI-anchor-depleted form, respectively. Both forms were adsorbed on dimyristoylphosphatidic acid LB films and restricted to the outermost layer. The surface density and enzyme activity were determined using a quartz crystal microbalance and p-nitrophenylphosphatase activity, respectively. The detergent-solubilized form was co-spread with dimyristoylphosphatidic acid on the air/water interface and transferred to solid supports, providing an enzyme maximum surface density of 530 ng/cm2. Maximal phosphohydrolytic activity, corresponding to 43% of that observed in homogeneous medium, was obtained at a surface density of 179 ng/cm2. The phospholipase C-solubilized form was adsorbed directly from solution, reaching a maximum surface density of 1541 ng/cm2, although the phosphomonohydrolase activity was 10 times lower than that obtained for the anchor-containing form. The combined analysis of surface density and enzymatic activity suggests that the alignment of the protein molecules on the LB lipid films induced by the glycosylphosphatidylinositol anchor facilitates the access of the substrate to the active site. This access is hampered by increasing enzyme surface densities and depends on a specific orientation of the adsorbed enzyme.