María Monteserín

Interaction between Poly(9,9-bis(6′-N,N,N-trimethylammonium)hexyl)fluorene phenylene) Bromide and DNA as Seen by Spectroscopy, Viscosity, and Conductivity: Effect of Molecular Weights and DNA Secondary Structure

The interaction between three poly(9,9-bis(6-N,N,N-trimethylammonium)hexyl)fluorene phenylene) bromide (HTMA-PFP) samples of different molecular weights (Mn=14.5, 30.1 and 61.3 kg/mol) and both dsDNA and ssDNA secondary structures has been studied using UV-visible absorption and fluorescence spectroscopies (including steady-state, time-resolved, and anisotropy measurements for the latter), viscosity, and electrical conductivity in 4% (v/v) DMSO-water mixtures. At low nucleic acid concentrations, formation of a 1:1 complex in terms of HTMA-PFP repeat units and DNA bases occurs. This interaction results in quenching of polymer emission. For higher molar ratios of DNA to HTMA-PFP, corresponding to charge neutralization, a second process is observed that is attributed to aggregate formation. From the changes in the absorption spectra, the polymer aggregation constant and the aggregate absorption spectra were calculated by applying an iterative method. Polymer aggregation dramatically quenches HTMA-PFP fluorescence in the region of the electroneutrality point. Under these conditions, the ratio of the emission intensity at 412 nm (maximum) to that at 434 nm (I412/I434) reaches a minimum, the electrical conductivity decreases, and the viscosity of the solution remains constant, showing that the DNA concentration can be determined through various HTMA-PFP physicochemical properties. With respect to the photophysical parameters (emission quantum yield, shape and shift of emission spectra), no significant differences were observed between dsDNA and ssDNA or with conjugated polymer or DNA molecular weight. The two short-lived components in the fluorescence decays are attributed to the presence of aggregates. Aggregates are also suggested to be responsible for the decrease in the fluorescence anisotropy through interchain exciton migration.

β-Phase Formation of Poly(9,9-dioctylfluorene) Induced by Liposome Phospholipid Bilayers

The well-structured β-phase emission of the neutral poly(9,9-dioctylfluorene) (PFO) is observed in 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayers, either as polydisperse aqueous liposomes or as the lamellar phase in thin films, and has been characterized by absorption, fluorescence (steady-state and time-resolved), and fluorescence anisotropy spectroscopy. Inclusion of PFO in DMPC liposomes provides a way of obtaining the ordered structure of this neutral polymer in aqueous suspensions. Quantification of the increase of the PFO β-phase in DMPC liposomes with the increase in polymer concentration is followed by deconvolution of the absorption spectra. In solid films, the presence of the phospholipids enhances the β-phase formation. In addition, the effect of the PFO concentration on the phospholipid phase transitions has been studied by differential scanning calorimetry (in liposome) and polarized light thermal microscopy (in solid film), confirming PFO/DMPC interactions in both liposome and films. The liposome size and structure in the presence and absence of polymer were characterized by dynamic light scattering and transmission electron microscopy, which showed relatively modest changes in liposome shape but a decrease in size upon incorporation of PFO.

Effects of the Interaction Between β-Carboline-3-carboxylic acid N-Methylamide and Polynucleotides on Singlet Oxygen Quantum Yield and DNA Oxidative Damage

The complexation of β‐carboline‐3‐carboxylic acid N‐methylamide (βCMAM) with the sodium salts of the nucleotides polyadenylic (Poly A), polycytidylic (Poly C), polyguanylic (Poly G), polythymidylic (Poly T) and polyuridylic (Poly U) acids, and with double stranded (dsDNA) and single stranded deoxyribonucleic acids (ssDNA) was studied at pH 4, 6 and 9. Predominant 1:1 complex formation is indicated from Job plots. Association constants were determined using the Benesi–Hildebrand equation. βCMAM‐sensitized singlet oxygen quantum yields were determined at pH 4, 6 and 9, and the effects on this of adding oligonucleotides, dsDNA and ssDNA were studied at the three pH values. With dsDNA, the effect on βCMAM triplet state formation was also determined through triplet–triplet transient absorption spectra. To evaluate possible oxidative damage of DNA following singlet oxygen βCMAM photosensitization, we used thiobarbituric acid‐reactivity assays and electrophoretic separation of DNA assays. The results showed no oxidative damage at the level of DNA degradation or strand break.

A Spectroscopic Study of the Interaction of the Fluorescent β-Carboline-3-carboxylic Acid N-methylamide with DNA Constituents: Nucleobases, Nucleosides and Nucleotides

Interaction between β-carboline-3-carboxylic acid N-methylamide, βCMAM, and nucleobases, nucleosides and nucleotides is studied in the ground state with UV-visible, 1H NMR and 31P NMR spectroscopies and in the first excited state, with steady-state and time-resolved fluorescence spectroscopy. Job plots show a predominant 1:1 interaction in both electronic states. Association constants are estimated from changes in the absorption spectra, and show that the strongest interaction is produced with the nucleosides: 2′-deoxyadenosine (dAdo) and thymidine (Thd), and with the mononucleotides: 2′-deoxycytidine 5′- monophosphate (5′-dCMP) and uridine 5′- monophosphate (5′-UMP). These results are corroborated by the upfield shifts of two 1H NMR resonances of the βCMAM indole group. The 31P NMR resonance of nucleotides is shifted downfield, suggesting the presence of electrostatic or hydrogen bond interaction with βCMAM. In the first electronic singlet excited state, static and dynamic quenching of βCMAM emission is achieved upon addition of nucleobases, nucleosides and nucleotides. This has been analysed using Stern–Volmer kinetics.

Effect of the Phospholipid Chain Length and Head Group on Beta‐Phase Formation of Poly(9,9‐dioctylfluorene) Enclosed in Liposomes

We have studied the effect of head group and alkyl chain length on β‐phase formation in poly(9,9‐dioctylfluorene) (PFO) solubilized in phospholipid liposomes. Systems studied have three different alkyl chain lengths (1,2‐dimyristoyl‐sn‐glycero‐3‐phosphatidylcholine [DMPC], 1,2‐didodecanoyl‐sn‐glycero‐3‐phosphatidylcholine [DLPC], 1,2‐dipalmitoyl‐sn‐glycero‐3‐phosphatidylcholine [DPPC]) and head groups (1,2‐dimyristoyl‐sn‐glycero‐3‐phosphate monosodium salt [DMPA], 1,2‐dimyristoyl‐sn‐glycero‐3‐phosphoethanolamine [DMPE] and 1,2‐dimyristoyl‐sn‐glycero‐3‐phospho‐l‐serine sodium salt [DMPS]). Changes in liposome size upon addition of PFO are followed by dynamic light scattering. All the phospholipids induce the formation of PFO β‐phase, which is followed by the emission intensity and deconvolution of the absorption spectra. Both the head group and alkyl chain length affect the yield of β‐phase. The photophysics of PFO incorporated in liposomes is characterized by stationary and time‐resolved fluorescence, whereas the polymer‐phospholipid interactions have been studied by the effect of the PFO concentration on the phospholipid phase transitions (differential scanning calorimetry [DSC]).