Journal of Photochemistry and Photobiology A: Chemistry 216 (2010) 79–84 Contents lists available at ScienceDirect Journal of Photochemistry and Photobiology A: Chemistry journal homepage: www.elsevier.com/locate/jphotochem Synthesis and spectral properties of fluorescent linear alkylphosphocholines labeled with all-(E)-1,6-diphenyl-1,3,5-hexatriene Valentín Hornillos a,b , Laura Tormo a , Francisco Amat-Guerri b , A. Ulises Acu ˜ na a,∗ a Instituto de Química Física Rocasolano, CSIC, Serrano 119, 28006 Madrid, Spain b Instituto de Química Orgánica General, CSIC, Juan de la Cierva 3, 20006 Madrid, Spain article info Article history: Received 16 July 2010 Received in revised form 6 September 2010 Accepted 8 September 2010 Available online 17 September 2010 Keywords: Diphenylhexatriene Lipid probes Fluorescence Synthesis Alkylphosphocholines abstract A synthetic method has been designed to produce alkylphosphatidylcholine lipid molecules with the fluorescent group all-(E)-1,6-diphenyl-1,3,5-hexatriene (DPH) incorporated to the end of a polymethy- lene chain of variable length. The resulting compounds may be viewed as single-chain fluorescent lipid probes terminated with a phosphocholine (PHC) polar head-group. The method was applied to the syn- thesis of two lipids with different alkyl chain length, PHC-C4-DPH (1) and PHC-C6-DPH (2), as well as the corresponding alcohols OH-C4-DPH (8) and OH-C6-DPH (9). The absorption and fluorescence properties of these compounds in fluid solution were very similar to those of the well-known lipid probe propionic acid-DPH (PA-DPH). In addition, it was observed that the alkylphosphocholines were easily incorporated into unilamellar lipid vesicles, in which the DPH group would be positioned most likely near the centre of the bilayer. In these conditions, 1 shows a fluorescence quantum yield of 0.4 ± 0.1 and a relatively simple decay time, with  av close to 5.0 ns. Fluorescence polarization experiments also indicate that the reorientational motions of the DPH group of phosphocholine 1 are sensitive to the fluidity changes that accompany the main thermal phase transition of the bilayer. As a result, both compounds may be of utility as novel fluorescent lipid probes, characterized by a biomimetic zwitterionic polar head-group. © 2010 Elsevier B.V. All rights reserved. 1. Introduction The study of lipid function in natural or artificial systems could be greatly simplified by means of the current large array of exper- imental methods based on fluorescence spectroscopy. However, since lipids usually lack intrinsic fluorescence, the application of these methods requires first attaching a suitable emitting group to the lipid molecule, either at its hydrophilic head-group or at the hydrophobic alkyl chain [1]. As a result of that, the delicate amphipatic balance of the original molecule is altered, frequently perturbing one or several properties of the parent compound. We were interested in the application of fluorescence microscopy techniques to understand the antiparasite mechanism of single- chain alkyl-lipids, as the phosphocholine (PHC) ester of the fatty alcohol n-hexadecanol (miltefosine, Fig. 1), a drug used to treat human leishmaniasis infections and other diseases [2]. The PHC head-group of the molecule is important for its bioactivity and, therefore, fluorescence labeling of the alkyl-lipid can only be car- ried out at the polymethylene chain. Simple conjugated polyenes, like tetraene [3,4] or pentaene [5,6] groups, may be inserted ∗ Corresponding author at: Instituto de Química Física Rocasolano, CSIC, Serrano 119, 28006 Madrid, Spain. Tel.: +34 91 7459501; fax: +34 91 5642431. E-mail address: roculises@iqfr.csic.es (A.U. Acu ˜ na). in the saturated alkyl chain as emitting labels, hopefully mini- mally perturbing the original physico-chemical properties of the drug [1]. Unfortunately, the UV excitation (ca. 350 nm) required by these weakly emitting groups is inconvenient for living-cell fluorescence imaging methods, although in some instances two- photon excitation techniques may overcome this limitation [7]. Conjugated phenyl-polyenes show red-shifted absorption tran- sitions and, in fact, a fluorescent analogue of miltefosine with potent antiparasite activity was produced by attaching a phenyl- tetraene chromophore to the parent drug [8]. Both, the excitation wavelength (ca. 380 nm) and the photochemical stability of this compound were more favorable than those of the simpler linear conjugated polyenes mentioned above. Further improvement in the photochemical and photophysical properties of the analogues may be achieved using all-(E)-1,6-diphenyl-1,3,5-hexatriene (DPH) as emitting tag. The lipophilic DPH fluorophore became popular in lipid mem- brane research because of its large absorption coefficient and fluorescence yield, extensive dynamic characterization in fluid solution and lipid bilayers and, importantly, high photochemical stability [9–11]. A series of lipid probes have been produced by attaching simple polar head-groups to the DPH fluorophore, such as the trimethylammonium cation [12–15] or the negatively charged propionic acid group [16–18]. In this way, the transverse location of the fluorescent tag of the probe in a lipid membrane can be better 1010-6030/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.jphotochem.2010.09.010