Spectral Properties of a Fluorescent Probe, all-trans-1,6-Diphenyl-1,3,5-hexatriene. Solvent and Temperature Effects

Abstract

Studying electronic absorption and fluorescence spectra of 1,6-diphenyl-1,3,5-hexatriene (DPH) in twenty-two solvents, several factors were found to affect the spectral parameters. Two groups of DPH spectral behaviour were defined, related to the refractive index and the dispersion interactions parameters of the solvents. DPH molar absorptivity is hardly affected by the nature of the solvent or the increase of temperature (20-50°C). By contrast, DPH fluorescence intensity increases along with the viscosity and polarizability of solvents and is decreased by elevating the temperature. Investigations on the combined effects of solvent nature and temperature on DPH spectral parameters allowed the solvents to be classed into four groups (referred to as 1 to 4). When the temperature was increased from 20 to 50°C, DPH fluorescence intensity was only slightly modified in solvents from group 1 (as tetrahydrofuran, I = 100 at 20°C, I = 80 ± 5 at 50°C), but was severely decreased in solvents from groups 2 (as ethanol, I = 55 at 50°C) and 3 (as acetic acid, I = 45 at 50°C), and still more in solvents from group 4 (tetrachloromethane, I = 10 at 50°C). Data on the reversibility of the phenomenon in these solvents after returning the temperature from 50 to 20°C show a concomitant decrease in DPH stability in these solvents. When a water-miscible solvent was used, the decrease in fluorescence intensity was found to be higher after adding water to this solvent. These data may help to assess the spectral behaviour of DPH in the more complex biological media.Fluorescent probes are extensively used in physical, chemical and biological sciences for investigating the structure and dynamics of living systems. 1 One of the most commonly used probes for the assessment of membrane microviscosity by fluorescence polarization is the polyene 1,6-diphenyl-1,3,5-hexatriene (DPH). Due to its long rod-like structure and its non-polar properties, DPH aligns parallel to the phospholipid fatty acyl chains of biological membranes. These properties explain its wide use for measuring the fluidity of membrane bilayers. 2 Yet, difficulties have been reported in using this probe, due to its poorly known location in the membranes, 3,4 its partition in cellular compartments, 5-7 and its fluorescence quenching caused by DPH-water interactions. 8 Unusual fluorescence properties of DPH have been listed 8 but only a few solvents, namely methylcyclohexane, heptane, benzene, ethyl iodide, perfluorohexane, acetonitrile, 1,4-dioxane and ethanol, have been used for studying the physicochemical properties of the probe. 9-11 Although excited states of DPH have been well studied because of its structural analogy with retinol, 12-15 the influence of the surrounding medium on the spectral properties of DPH is not yet clear. In the present study, the interactions between DPH and various solvents were examined; it is shown that the stability of the probe largely depends on the temperature and the nature or hydration of solvents. ExperimentalTwenty-two solvents of spectrop...

Unusual 1,6-diphenyl-1,3,5-hexatriene (DPH) spectrophotometric behavior in water/ethanol and water/DMSO mixtures

“…19 This effect of water on the DPH fluorescence was previously reported by Dupuy and Montagu. 2 In that work the fluorescence intensity decayed 16 and 29% in ethanol, and 3 and 13% in DMSO for 3 and 10% water, respectively. They assigned this phenomenon to DPH and water interactions without further discussion.…”

“…9,15 However, the large decrease in the fluorescence intensity for such low water percentage in the solvent cannot be attributed to a simple solvent polarizability effect, as demonstrated by the fluorescence intensity decay of 27% for the addition of 10% water in methanol. 2 The a S of methanol is even smaller than that of water (a S, methanol = 0.203). As the same time the observed fluorescence decrease can not be attributed to dielectric constant effects.…”

“…1,2 The spectrophotometric properties of DPH are very sensitive to the experimental conditions, which allows using this compound and its derivatives as environmental probes in colloidal systems to determine the critical micellar concentration of surfactants 3 and in biological cell membranes. 4,5 The measurement of the fluorescence anisotropy of DPH is extensively used for membranes, particularly in studies of microviscosities, 6 due to the non-polar characteristic, the low water solubility, and the suitable geometric design, which maintains the molecule aligned parallel to the phospholipids chains of the membranes.…”

“…7 Although the DPH molecule remains inserted in the membrane layer, it stays in contact with water molecules. 2,5 The majority of DPH spectrophotometric studies on electronic transitions are performed in organic solvents. 18 In fact, either fluorescence or rotation (cis-trans isomerization) occurs from 1 A g * (S 1 state).…”

Influence of the N- and C-Terminal Regions of Antimicrobial Peptide Pleurocidin on Antibacterial Activity

“…To investigate biological properties of the terminal regions, Anal 1 and Anal 2 (7-25) were designed by cutting GWG (1-3) or GWGSFF (1-6) residues in the N-terminal region, respectively. In addition, we synthesized Anal 3 and Anal 4 (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19), which are truncated peptides of pleurocidin with HYL (23)(24)(25) or ALTHYL (20)(21)(22)(23)(24)(25) in C-terminal deletion.…”