Computational Study on Cycloisomerization/Oxidative Dimerization of Aryl Propargyl Ethers Catalyzed by Gold Nanoclusters: Mechanism and Selectivity

A theoretical analysis of the cycloisomerization and oxidative dimerization of phenyl propargyl ether catalyzed by the Au₃₈ cluster is performed by means of density functional theory. The role of the cationic gold species is also clarified. The substituent effect at the para site of phenyl is studied to explore the selectivity of cycloisomerization and oxidative dimerization. Phenyl propargyl ether preferred to adsorb at the T1 site of the Au₃₈ surface with an adsorption energy of −10.61 kcal/mol. The 6-<i>endo</i> pathway to give 2<i>H</i>-chromene is the most feasible pathway, with an energy barrier of 20.50 kcal/mol in dichloroethane solvent. The energy barriers of the 5-<i>exo</i> and oxidative dimerization pathways in dichloroethane solvent are 25.81 and 30.14 kcal/mol, respectively. 2<i>H</i>-Chromene is the main product of the cycloisomerization of phenyl propargyl ether catalyzed by the gold cluster. The presence of the cationic gold species can increase the yield of dimeric 2<i>H</i>-chromene, which is in agreement with experiment results. The binding strength between the active sites and 2<i>H</i>-chromen-3-yl is crucial for oxidative dimerization. Substituents at the para site of phenyl have only a slight influence on the 6-<i>endo</i> pathway, except for the methoxyl group. The differences in the energy barriers between cycloisomerization and oxidative dimerization are in agreement with the ratio of 2<i>H</i>-chromenes and 2<i>H</i>,2′<i>H</i>-4,4′-bichromenes obtained in experiments. The selectivity for the 6-<i>endo</i>/dimeric pathways is sensitive to the substituents of the substrates and the electronic prosperities of the active site of the catalysts. Our theoretical results are in agreement with the product distribution and phenomena in experiments

miR21 is directly regulated by NF-kB.

<p>Real-time PCR results for miR-21 levels were performed with siRNA against NF-κB. After the transfection of siRNA against NF-κB, the miR-21 level was detected by real-time PCR (6A). **<i>P</i><0.01, the siRNA treatment group vs the non- siRNA treatment group. Then, the apoptosis level was analyzed by flow cytometry (6D). **<i>P</i><0.01, the siRNA treatment group vs the non- siRNA treatment group. Five primers were designed according to a software analysis, and ChIP assays were performed in RAW264.7 cells to explore possible binding sites. A specific band was observed for primer for site 3 (6B). No specific band was observed for the primers for sites 1, 2, 4 and 5 (6C). Negative controls were incubated without primary antibody. The experiment was repeated three times.</p

Alignment of the combined plastid dataset

Alignment of the combined plastid datase

Bayesian tree of the tribe-wide nrITS dataset

Bayesian tree of the tribe-wide nrITS datase

Maximum Likelihood tree of the nuclear Xdh dataset

Maximum Likelihood tree of the nuclear Xdh datase

Alignment of the tribe-wide nrITS dataset

Alignment of the tribe-wide nrITS datase

Maximum Likelihood tree of the tribe-wide nrITS dataset

Maximum Likelihood tree of the tribe-wide nrITS datase

The control effect of miRNA21 on bcl-2 after MPT64 treatment RAW264.7.

<p>Macrophages were treated as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100949#pone-0100949-g002" target="_blank">Fig. 2</a>. The miRNA21 level was analyzed by real time-PCR (5A). **<i>P</i><0.01, the PPD group vs the PPD-MPT64 (15 µg/ml) group. The control effect of miRNA21 on bcl-2 was also analyzed by relative luciferase activity in RAW264.7 cells (5B). The wild-type or mutated type of bcl-2-utr was cloned into a pMIR-reportTM luciferase vector. Then, the recombinant plasmids and miR21 were transfected into RAW264.7 cells. The relative luciferase activity was detected. miR-mock and miR335 were used as control miRNAs. *<i>P</i><0.01, the miR21 group vs the miR-mock group after the transfection by wild-type bcl-2-utr. Data shown are representative of three independent experiments.</p

Apoptosis inhibition of macrophages by MPT64 protein.

<p>PMA-differentiated RAW264.7 cells were put into 24-well flat bottom tissue culture plates at a density of 1×10⁵ cells per well. Then, RAW264.7 macrophages were incubated with PBS, BCG-PPD (10 µg/ml) or a mixture of PPD-MPT64 at a different concentrations for 16 h. (A) Apoptosis was detected by measuring the membrane exposure of PS using annexin V by flow cytometry, and the results were analyzed. <b>Fig. 2A</b> Significant differences of the apoptosis percentages. *<i>P</i><0.05, the PPD group vs the MPT64 (10 µg/ml) group; ** <i>P</i><0.01, the PBS group vs the PPD group, and the PPD group vs the MPT64 (15 µg/ml or 20 µg/ml) group. The apoptosis percentage is not significantly different for the GST group or heat-treated MPT64 protein compared with the PPD group. <b>Fig. 2A</b> Ne, the PBS group; HAMPT64, heat treated MPT64; M10, MPT64 at 10 µg/ml of concentration; M15, MPT64 at 15 µg/ml of concentration; M20, MPT64 at 20 µg/ml of concentration. <b>Fig. 2B</b> A: The PBS group. B: The PPD group. C: The MPT64 (15 µg/ml) group. Data shown are representative of three independent experiments.</p

Maximum Parsimony strict consensus tree of the combined plastid dataset

Maximum Parsimony strict consensus tree of the combined plastid datase