A fluorescent phospholipid derivative, the fluoresceinthiocarbamyl adduct of a natural phosphatidylethanolamine, has been synthesized and incorporated into sonicated single-bilayer vesicles of egg lecithin and dipalmitoyllecithin. The surface location of this probe has been confirmed by using extrinsic fluorescence quenching studies together with steady-state emission anisotropy measurements. Electronic excitation energy transfer between 1,6-diphenyl-1,3,5-hexatriene incorporated within the hydrophobic core of the bilayer and the novel derivative has been investigated to estimate the depth within the bilayer at which the former is located. Efficiencies have been measured for two different phospholipids, egg lecithin and dipalmitoyllecithin, in the latter case both above and below the phospholipid phase transition, with and without added cholesterol. The observed dependence of the transfer efficiency on the acceptor concentration was compared with that calculated according to Förster theory applied to random two-dimensional distributions of donor and acceptor molecules in parallel planes for various interplanar separations, taking into account orientational effects. The Förster R0 of about 45 A for this donor-acceptor pair is particularly well suited to such studies since it is of the order of the width of the bilayer. The experiments showed that energy-transfer spectroscopy can provide useful quantitative information as to the transverse location of diphenylhexatriene in homogeneous phospholipid bilayers and may also reflect lateral partitioning of donor or of both donor and acceptor into different phases in systems exhibiting phase separations.

The pH dependence of singlet oxygen quenching by histidine, N-acetyltyrosine ethyl ester (ATEE), ascorbic acid, Trolox C, and tryptophan has been observed using time-resolved infrared luminescence measurements in a D2O/acetonitrile (50:50 v/v) solvent. Deprotonation of ascorbic acid, the protonated imidazole ring of histidine and the phenolic group of ATEE leads to an increase in the quenching rate constants by between 2 and 3 orders of magnitude. Such changes appear to be the basis for wide variations in quoted literature values of singlet oxygen quenching constants for these and related compounds. It is estimated that these pH-dependent quenching rate constants predict a modest (approximately 2- to 3-fold) change in singlet oxygen lifetime between the extremes of cellular pH. Activation data for singlet oxygen quenching show that the enthalpies of activation are low in all cases (between 0 and 11 kJ mol-1) and that substantially negative entropies of activation (between −49 and −116 J K-1 mol-1) result in rate constants being much lower than the diffusion-controlled limit. In all cases the data are consistent with quenching via reversible formation of an exciplex, all reactions being at the preequilibrium limit over the available temperature range.

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Laser flash photolysis of α‐tocopherol in methanol and in aqueous micellar solutions has been shown to produce the α‐tocopheroxyl radical. The reaction between the α‐tocopheroxyl radical and ascorbate in positively charged hexadecyltrimethylammonium chloride (HTAC) micelles occurred with a second order rate constant of 7.2 × 107 M−1·s−1, whereas in negatively charged sodium dodecyl sulphate (SDS) micelles the rats constant was only 3.8 × 104 M−1·s−1. The α‐tocopheroxyl radical was found to be relatively long‐lived in HTAC micelles (t½ ⩾ 5 min), allowing the slow disappearance of the α‐tocopheroxyl radical by reaction with glutathione to be observed.

The real-time uptake of serotonin, a neurotransmitter, by rat leukemia mast cell line RBL-2H3 and 5-hydroxytryptophan by Chinese hamster V79 cells has been studied by fluorescence lifetime imaging microscopy (FLIM), monitoring ultraviolet (340 nm) fluorescence induced by two-photon subpicosecond 630 nm excitation. Comparison with two-photon excitation with 590 nm photons or by three-photon excitation at 740 nm shows that the use of 630 nm excitation provides optimal signal intensity and lowered background from auto-fluorescence of other cellular components. In intact cells, we observe using FLIM three distinct fluorescence lifetimes of serotonin and 5-hydroxytryptophan according to location. The normal fluorescence lifetimes of both serotonin (3.8 ns) and 5-hydroxytryptophan (3.5 ns) in solution are reduced to approximately 2.5 ns immediately on uptake into the cell cytosol. The lifetime of internalized serotonin in RBL-2H3 cells is further reduced to approximately 2.0 ns when stored within secretory vesicles.