Fluorescent probes for asymmetric lipid bilayers: Synthesis and properties in phosphatidyl choline liposomes and erythrocyte membranes

Abstract

We have synthesized three sets of fluorescent probes which we believe will be useful in studies of asymmetric membranes and have studied their interactions with model lipid bilayers and erythrocyte membranes. The probes were designed to partition preferentially into one face of a lipid bilayer with asymmetrically disposed phospholipids and to report lipid transitions in that monolayer. We synthesized more than twenty probes containing anthroyl-, dansyl-, or pyrene rings with acidic, basic, and neutral functional groups and alkyl "spacers" of various lengths. The interactions of these probes with liposomes of phosphatidyl choline and with erythrocyte membranes were characterized to determine whether probe insertion was asymmetric, how deeply the probe penetrated the bilayer, and whether the probe reflected thermotropic phase transitions in model membranes. The set of variously charged anthroyl esters, analogs of local anaesthetics, appears to be promising for studies of asymmetric membranes. Fluorescent probes have been used extensively to provide information on the lipid regions of biological membranes. Membrane fluidity, a composite of molecular packing and motion of acyl chains in lipid bilayers, has been assessed with a variety of fluorescent probes, the fluorescence of which undergoes some measurable change at the temperature of the membrane's thermotropic phase transition. A large number of fluorescent probes have been used for this purpose. Bashford, Morgan and Radda (Bashford, C.L., Morgan, C.G., Radda, G.K. 1976; Biochim. Biophys. Acta 426:157) and Thulborn and Sawyer (Thulborn, K.R., Sawyer, W. H. 1978; Biochim. Biophys. Acta 511:125) synthesized several fatty acid derivatives in which an anthracene group is attached (in ester linkage) along the acyl chain at various positions, and have shown that this set of probes may be useful in probing membrane fluidity at different depths within the bilayer. This report describes the synthesis and properties of several sets of amphipathic fluorescent probes, which may partition unequally into the two faces of an asymmetric lipid bilayer, and may therefore provide information about membranes complementary to that obtainable with existing probes.

Groups with Polar Characteristics Can Locate at Both Shallow and Deep Locations in Membranes: The Behavior of Dansyl and Related Probes

“…λ max values in model membranes relative to that in solvent of varying polarity. Such studies have indicated that dansyl and related derivatives are located somewhere in the polar headgroup region (1,31) in agreement with our results.…”

Synthesis of α,ω-Dimercapto Poly(N-isopropylacrylamides) by RAFT Polymerization with a Hydrophilic Difunctional Chain Transfer Agent

“…To determine c IPC , the pyrene-substituted initiator BPAC was used instead of AIBN to initiate the polymerization under the same conditions. The polymers obtained were aminolyzed to remove the thiocarbonylthio end groups, thus allowing precise determination of the content of pyrenyl end groups, using the absorbance at 342 nm and a molar extinction coefficient of 2 × ε 342 (ε 342,MeOH = 32 800 M -1 cm -1 for 4-(1-pyrene)butylamine chloride employed as model compound) . Two such samples, namely α,ω-SH-PNIPAM(py)-7k and α,ω-SH-PNIPAM(py)-11k (Table ), were prepared with polymerization times of 3 and 4 h. The c IPC values determined experimentally for polymers prepared with BPAC were also used to determine the M n value of polymers prepared with AIBN of similar polymerization time.…”

Synthesis of a Polymer-Supported Anthracene and Its Application as a Dienophile Scavenger

“…Our initial synthesis of a polymer-supported anthracene 2 is shown in Scheme 1. Acylation of 9-anthracenemethanol with succinic anhydride afforded anthracene acid 1 , which was coupled to commercially available aminomethyl polystyrene resin (PS−NH 2 , Argonaut Technologies, 1.52 mmol/g) to provide polymer-bound anthracene 2 . The scavenging capacity of 2 was determined by thermolysis with N -(4-bromophenyl)maleimide (3.0 equiv) in toluene at 70 °C (24 h).…”