Effects of C5-substituent group on the hydrogen peroxide-mediated tautomerisation of protonated cytosine: a theoretical perspective

<p>The direct tautomerism (path A) and H₂O₂ as a catalyst (path B) have been studied in conversion of Cyt2t⁺ into CytN3⁺ isomer. The protonated 5-carboxycytosine (5-caCyt) is represented and has been further explored in the presence of H₂O₂ (path C). In going from a four-membered-ring transition state in the case of the direct tautomerism to the six-membered ring for H₂O₂, the H₂O₂ significantly contributes to decreasing the free energy barrier of tautomerisation. Although the carboxylic substituent of 5-carboxycytosine has certain affected on the electron distribution of the pyrimidine ring, the six-membered-ring transition state has not changed. This result illustrates that the C5-carboxylation has no significant effect on the H₂O₂-mediated isomerisation of Cyt2t⁺ to CytN3⁺ isomer. Meanwhile, these paths A–C have been further explored in the presence of two water molecules. Use of implicit solvent models (PCM) does not significantly alter the energetics of water-mediated paths A–C compared to those in gas phase. Furthermore, the rate constant with Wigner tunnelling correction of path A is obviously smaller than those of paths B and C. Finally, the lifetime τ_99.9% of paths B and C is 10⁻⁵ s, which is implemented by the mechanism of the concerted synchronous double proton transfer.</p

Roemerine-mediated bacterial membrane permeabilization (original magnification, ×1000).

<p>(A) Cultured <i>S</i>. <i>aureus</i> ATCC29213 was treated with 0, 32, 64 or 128 μg/mL of roemerine for 6 h, then were stained with the dyes SYTO 9 (which stains live bacteria) and propidium iodide (which stains dead bacteria). The stained bacteria were analyzed by fluorescence microscopy to assess viability. (B) The numbers indicate the percentages of live bacteria within <i>S</i>. <i>aureus</i> ATCC29213 after treatment with different concentrations of Roemerine. (C) The numbers indicate the percentages of dead bacteria within <i>S</i>. <i>aureus</i> ATCC29213 after treatment with different concentrations of Roemerine.</p

Activity of roemerine against MRSA XJ75302 in a mouse sepsis model.

<p>Survival of BALB/c mice inoculated by i.p. injection with MRSA XJ75302 and treated with 20 mg/kg roemerine, oxacillin, vancomycin or sterile water by i.p. administration. Values with statistical significance by the log-rank test are indicated by an asterisks: *<i>P</i><0.05, ***<i>P</i><0.001 versus Model group.</p

Concentration-dependent inhibition of roemerine on the growth of four <i>S</i>. <i>aureus</i>.

<p>Roemerine was added to cell cultures containing different tested strains to a final concentration 8, 16, 32, or 64 μg/mL, with addition of an equal volume of sterile water as a control. The growth curves for four tested strains were measured using a BioscreenC™ instrument in the absence and presence of different concentrations of roemerine. The sample frequency was one hour and data at time points are the means for three replicates. ROE, roemerine. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001 versus Control.</p

Effects of roemerine on the growth of bacteria colonies.

<p>Roemerine, vancomycin and oxacillin were added to cell cultures containing <i>S</i>. <i>aureus</i> ATCC29213, MRSA XJ75302, Mu50, or LAC, respectively, with addition of an equal volume of sterile water as a control. Aliquots of each culture were collected at 0, 3, 6, 12, and 24 h, and were then diluted and inoculated on solid agar. The number of colony-forming units (CFUs) was calculated from the number of colonies growing on plates, and the data are the means for three replicates. CON, control; ROE, roemerine; VAN, vancomycin; OXA, oxacillin. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001 versus Control.</p

Using Gold Nanoclusters As Selective Luminescent Probes for Phosphate-Containing Metabolites

Glutathione-bound gold nanoclusters (AuNCs@GSH) can emit reddish photoluminescence under illumination of ultraviolet light. The luminescence of the AuNCs@GSH is quenched when chelating with iron ions (AuNCs@GSH-Fe³⁺), presumably resulting from the effective electron transfer between the nanoclusters and iron ions. Nevertheless, we found that the luminescence of the gold nanoclusters can be restored in the presence of phosphate-containing molecules, which suggested the possibility of using AuNCs@GSH-Fe³⁺ complexes as the selective luminescent switches for phosphate-containing metabolites. Phosphate-containing metabolites such as adenosine-5′-triphosphate (ATP) and pyrophosphate play an important role in biological systems. In this study, we demonstrated that the luminescence of the AuNCs@GSH-Fe³⁺ is switched-on when mixing with ATP and pyrophosphate, which can readily be observed by the naked eye. It results from the high formation constants between phosphates and iron ions. When employing fluorescence spectroscopy as the detection tool, quantitative analysis for phosphate-containing metabolites such as ATP and pyrophosphate can be conducted. The linear range for ATP and pyrophosphate is 50 μM to sub-millimolar, while the limit of detection for ATP and pyrophosphate are ∼43 and ∼28 μM, respectively. Additionally, we demonstrated that the luminescence of the AuNCs@GSH-Fe³⁺ can also be turned on in the presence of phosphate-containing metabolites from cell lysates and blood plasma