Protonation of porphyrins in liquid PVC membranes: Effects of anionic additives and application to pH-sensing

We report on a study on the protonation of representative porphyrins, metalloporphyrins, porphyrin-ketones and a Pd-porphyrin Schiff base in aqueous solution and when dissolved in plasticized PVC membranes. The respective protolytic forms were characterized by absorption and emission spectra as well as apparent pKa values. It is shown that the protonation of porphyrins dissolved in PVC membranes is much more difficult than in solution, resulting in an extraordinarily large decrease in the apparent pKa values which can be as large as 5 units when compared to the corresponding data for the porphyrins in solution. In certain cases, no protonation at all occurred within the pH 2–12 range. At the same time, N-MeEP which is much more basic (its intrinsic pK being 11.2) can be protonated in such membranes into the monocation at weakly acidic pH, but the apparent pKa also is lowered by about 6 units. On addition of a tetraphenylborate anion to the PVC membranes, protonation was found to proceed much easier. Both the monocation and dication was identified in case for the porphyrins, while for the prophyrin-ketones protonation into dication took place, and N-methyl-etioprophyrin was present as the monocation over the whole pH 2–12 range. Membranes composed of solutions of porphyrin-ketones or the Pd-porphyrin Schiff base in plasticized PVC were identified as being useful for purposes of continuously sensing pH in the physiological range. Their response time, calibration curves, signal stability and effects of buffer ionic strength and temperature were investigated in more detail

Longwave luminescent porphyrin probes

A set of luminescent dyes, namely the porphyrin-ketones and their zinc(II), platinum(II) and palladium(II) complexes, are described. The probes are derived from a partially oxidized porphyrin ring and have been studied by absorption and luminescence spectroscopy including time-resolved measurements. The metal-free porphyrin-ketones display strong and pH-dependent fluorescence due to a protolytic equilibrium that exists between the free base and the dication. The zinc(II) complexes also give strong fluorescence along with a weak long-decay emission at room temperature which are attributed to delayed fluorescence and phosphorescence, respectively. Platinum(II) and palladium(II) complexes exhibit room-temperature phosphorescence without any detectable fluorescence. Compared to existing porphyrins, the new dyes display a considerably longwave-shifted luminescence along with substantially improved (photo)chemical stability. They are considered to be promising probes for very near-infrared luminescence studies, and to be particularly useful for applications such as labelling of biomolecules, pH-sensing, and detection of quenching species such as molecular oxygen

Cell-penetrating conjugates of coproporphyrins with oligoarginine peptides : rational design and application for sensing intracellular O-2

A panel of phosphorescent oligoarginine conjugates of tetracarboxylic Pt(II)-coproporphyrin I dye (PtCP), monosubstituted with long peptides or tetra-substituted with short peptides and having different linkers and peripheral groups, is described. Their photophysical properties, cell loading efficiency, and mechanisms of transport into the cell were investigated and compared. The conjugates were seen to rely on endocytotic mechanisms of cell entry, which are different from that of the unconjugated oligoarginine peptide, and show diverse patterns of intracellular distribution. On the basis of this study, the tetra-substituted PtCP conjugate displaying whole cell distribution was selected for the sensing of intracellular O-2. This probe has been tested in biological experiments on a fluorescence plate reader, including the monitoring of in situ oxygenation of respiring cells and their responses to metabolic stimulation. Similar conjugates of the phosphorescent Pd(II)-coprorphyrin and fluorescent coproporphyrin-ketone were also synthesized and assessed for the sensing of low levels intracellular O-2 and ratiometric pH-sensing, respectively. The results produced and the structure-activity relationships determined can facilitate the rational design of new bioconjugates of porphyrin dyes tailored to specific applications

Synthesis, Solution, Molecular and Crystal Structure of Bis[5-(coproporphyrinato-I tetraethylester)nickel(II)]1,2-ethane

Vilsmeier formylation of the oporphyrin-I tetraethyl ester Ni-complex (III) followed by reduction of the intermediary imine salt (IV) with NaBH4 gave the o-dimethylaminomethyl-coproporphyrin-I Ni-complex (V). Heating the latter in methyl iodide gave 65% of the ethane-bis-porphyrin (I), which has been characterized by NMR-spectroscopy and a molecular and crystal structure investigation. The crystal structure shows an extended structure of the bisporphyrin with a parallel arrangement of the porphyrin subunits. Crystal data: I -C90H106N8Ni2O16. Triclinic, a = 12.319(3) Å, b = 13.297(3) Å, c = 14.444(3) Å, α = 105.56(2)°, β = 100.82(2)°, γ = 112.98(2)°, V = 1981.0(7) Å3, Cu Kα radiation, λ = 1.54178 Å, space group P1, Z = 1, R = 0.07

Bactenecin 7 peptide fragment as a tool for intracellular delivery of a phosphorescent oxygen sensor

Research on cell-penetrating peptides for the intracellular delivery of porphyrin compounds has mainly focused on the use of trans-activator of transcription (TAT)-derived peptides and, to a lesser extent, on proline-rich peptides and phosphorescent metalloporphyrins. In this article, we describe a novel phosphorescent oxygen-sensitive probe for intracellular use which comprises a bactenecin 7 peptide fragment (15-24) conjugated with the uncharged monofunctional derivative of Pt(II) coproporphyrin I (PEPP0). This probe provides efficient loading of various mammalian cells, including PC12, HCT116, SH-SY5Y and HeLa, via cell-type-dependent uptake mechanisms. The conjugate displays a similar distribution in cytoplasm and mitochondria which allows local oxygen levels to be monitored. Respiratory responses of PC12 cells loaded with the conjugate, measured on a time-resolved fluorescent reader, showed significant cell deoxygenation in response to uncoupling by carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone and external hypoxia. Treatment with mitochondrial inhibitors led to a decrease in cell deoxygenation. Although the biophysical properties of this conjugate are similar to those of the phosphorescent intracellular oxygen-sensitive probes described previously, it possesses a number of advantages, including ease of synthesis, high loading efficiency and reliability in physiological experiments with cells. This intracellular probe can be employed for the measurement of intracellular O(2) levels in samples containing mammalian cells using the phosphorescence quenching technique. In addition, the responses to metabolic stimuli can be assessed in a wide range of cells, as can the levels of relative cell oxygenation under external hypoxia