Highly Fluorescent Distyrylnaphthalene Derivatives as a Tool for Visualization of Cellular Membranes

Fluorescent imaging, which is an important interdisciplinary field bridging research from organic chemistry, biochemistry and cell biology has been applied for multi-dimensional detection, visualization and characterization of biological structures and processes. Especially valuable is the possibility to monitor cellular processes in real time using fluorescent probes. In this work, conjugated oligoelectrolytes and neutral derivatives with the distyrylnaphthalene core (SN-COEs) were designed, synthetized and tested for biological properties as membrane-specific fluorescent dyes for the visualization of membrane-dependent cellular processes. The group of tested compounds includes newly synthesized distyrylnaphthalene derivatives (DSNNs): a trimethylammonium derivative (DSNN-NMe3+), a phosphonate derivative (DSNN-P), a morpholine derivative (DSNN-Mor), a dihydroxyethylamine derivative (DSNN-DEA), a phosphonate potassium salt (DSNN-POK), an amino derivative (DSNN-NH2) and pyridinium derivative (DSNN-Py+). All compounds were tested for their biological properties, including cytotoxicity and staining efficiency towards mammalian cells. The fluorescence intensity of SN-COEs incorporated into cellular structures was analyzed by fluorescence activated cell sorting (FACS) and photoluminescence spectroscopy. The cytotoxicity results have shown that all tested SN-COEs can be safely used in the human and animal cell studies. Fluorescence and confocal microscopy observations confirm that tested COEs can be applied as fluorescent probes for the visualization of intracellular membrane components in a wide range of different cell types, including adherent and suspension cells. The staining procedure may be performed under both serum free and complete medium conditions. The presented studies have revealed the interesting biological properties of SN-COEs and confirmed their applicability as dyes for staining the membranous structures of eukaryotic cells, which may be useful for visualization of wide range of biological processes dependent of the extra-/intracellular communications and/or based on the remodeling of cellular membranes

Vicinal diphosphoniums: electrostatic repulsion under covalent constraint

A series of five vicinal bis(alkyl-triarylphosphoniums) derived from o-bis(diphenyl-phosphino)benzene (o-dppb) is described. Each of them have been prepared by specific methods, and their formal electrostatic and possible Van der Waals strain is compared through the P+...P+ distances in the crystal state. According to X-ray diffraction analyses, while the conformations of the dimethyl dication is C2 symmetric, the conformation of alka-1,n-diyl-diphosphoniums (n = 1, 2, 3) is pseudo-Cs symmetric. The solution structure of the dimethyldiphosphonium 2 was studied by NMR techniques after metathesis of the triflate counterions with enantiomerically pure X-PHAT chiral anions (X = TRIS, BIN). [(Delta)-BINPHAT] was indirectly shown to discriminate between the enantiomers of the diphosphonium, but no effective enantio-differentiation could be measured down to -70 degrees C in CD2Cl2. When associated to [(Delta)-TRISPHAT] counterions, progressive cleavage of the diphosphonium afforded the corresponding methyl-triphenyl-monophosphonium salt. In the bridged series, the diphosphoniacyclopentene dication was found to be stable in spite of the very short transannular P+...P+ distance (2.83 A). The naturally more relaxed homologous diphosphoniacyclohexene dication (P+...P+ = 3.28 A) was found to be flexible in solution, as indicated by the equivalence of the four protons of the +P-CH2CH2-P+ bridge from 20 degrees C to -80 degrees C in acetone-d6. The two diphosphoniacycloheptene dications exhibit no P+...P+ steric compression (ca 3.64 A), just as the dimethyl dication (3.70 A). In solution, the seven-membered ring of the +P-CH2CR2CH2-P+ dications (R = H, Me) remains rigidly Cs symmetric: no interconversion of the pseudo-axial and pseudo-equatorial R groups is evidenced at the NMR time scale at -95 degrees C (coalescence at -20 degrees C for R = H, at -65 degrees C for R = Me). According to DFT calculations at the B3PW91/6-31G** level in 1,2-dichloroethane (DCE: PCM, epsilon = 10.36), the lowest energy conformation is indeed Cs-symmetric. However a quasi-isoenergetic C2-symmetric conformation which is the most stable in the gas phase is also found, and is thus a likely intermediate in the observed Cs Cs interconversion. This +P-CH2CMe2CH2-P+ diphosphonium was obtained by dialkylation of o-dppb with 2,2-dimethylpropan-1,3-diyl bistriflate in DCE at 80 degrees C. The use of this solvent was the key for the synthesis of this dication, which was however competitively produced along with some +P-CH2CH2-P+ dication. The latter was also obtained by a totally different route, by heating a sulfinylethyl monophosphonium salt of o-dppb in the presence of the cationic complex [Rh(cod)2][PF6]. A mechanism for this peculiar process is proposed