Polyazamacrocycles based on a tetraaminoacetate moiety and a (poly)pyridine intracyclic unit: direct synthesis and application to the photosensitization of Eu(III) and Tb(III) ions in aqueous solutions

A series of five new 15-, 18- or 21-membered polyazamacrocycles (L₁-L₅) based on a pyridine, bipyridine or terpyridine unit and a triethylenetetraminetetraacetic acid (TTTA) skeleton is described. In ligands L₄ and L₅ the azaheterocycle contains an additional extracyclic functionality (ester group) suitable for covalently attachment to bioactive molecules. The synthetic procedure is based on the use of a linear tetra-N-alkylated tetramine synthon incorporating masked acetate arms and an efficient metal template ion effect, which controls the crucial macrocyclization step. In the case of L₁-L₃, the formation of lanthanide complexes with europium(III) and terbium(III) was investigated and the fluorescence characteristics of the complexes were established. In this series, the terbium(III) complex derived from the bipyridine ligand exhibits the highest lifetime and quantum yield values (τ=2.18 ms, Φ=26%).     

New Components Including Cyclopeptides from Barks of Christiana africana DC. (Tiliaceae)

Phytochemical investigation of barks of Christiana africana led to the identification of cyclopeptide alkaloids, flavonoids, coumarinolignans, iridoids, sesquiterpenoids, and triterpenes. This plant was classified so far in the Tiliaceae family. This study was started while the genomic study of numerous specimens was described in order to establish new criteria for Malvales botanical classification. In the present work, twenty components were identified, belonging to the three major classes of secondary metabolites: alkaloids, phenols, and terpenes. In the first class, cyclopeptides are well‐known compounds in Rhamnaceae and Sterculiaceae. Their presence in Malvaceae (in APG2 sensus) suggests a possible chemical link between the ex‐Tiliaceae and the Malvaceae.     

Immunochemical-based method for detection of hazelnut proteins in processed foods

A competitive enzyme-linked immunosorbent assay (ELISA) was developed to detect hazelnut by using polyclonal antibodies generated against a protein extract of roasted hazelnut. No cross-reactivity was observed in tests against 39 commodities, including many common allergens, tree nuts, and legumes. Hazelnut protein standard solutions at 0.45 ng/mL [inhibition concentration (IC80) of the competitive test] were clearly identified by the ELISA. An extraction and quantification method was developed and optimized for chocolate, cookies, breakfast cereals, and ice cream, major food commodities likely to be cross-contaminated with undeclared hazelnut during food processing. No sample cleanup was required when extracts were diluted 10-fold. Recovery results were generated with blank matrixes spiked at 4 levels from 1 to 10 microg/g hazelnut protein. With the developed extraction and sample handling procedure, hazelnut proteins were recovered at 64-83% from chocolate and at 78-97% from other matrixes. A confirmatory technique was developed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western transfer. The developed methods were applied to a small market survey of chocolate products and allowed the identification of undeclared hazelnut in these products.     

On the gene delivery efficacies of pH-sensitive cationic lipids via endosomal protonation: a chemical biology investigation

In an effort to probe the importance of endosomal protonation in pH-sensitive, cationic, lipid-mediated, non-viral gene delivery, we have designed and synthesized a novel cholesterol-based, endosomal pH-sensitive, histidylated, cationic amphiphile (lipid 1), its less pH-sensitive counterpart with an electron-deficient, tosylated histidine head group (lipid 2) as well as a third new cholesterol-based, cationic lipid containing no histidine head group (lipid 3). For all the novel liposomes and lipoplexes, we evaluated hysicochemical characteristics, including lipid:DNA interactions, global surface charge, and sizes. As anticipated, lipid 2 showed lower efficacies than lipid 1 for the transfection of 293T7 cells with the cytoplasmic gene expression vector pT7Luc at lipid:DNA mole ratios of 3.6:1 and 1.8:1; both lipids were greatly inhibited in the presence of Bafilomycin A1. This demonstrates the involvement of imidazole ring protonation in the endosomal escape of DNA. Conversely, endosome escape of DNA with lipid 3 seemed to be independent of endosome acidification. However, with nuclear gene expression systems in 293T7, HepG2, and HeLa cells, the transfection efficacies of lipid 2 at a lipid:DNA mole ratio of 3.6:1 were found to be either equal to or somewhat lower than those of lipids 1 and 3. Interestingly, at a lipid:DNA mole ratio of 1.8:1, lipids 2 and 3 were remarkably more transfection efficient than lipid 1 in both HepG2 and HeLa cells. Mechanistic implications of such contrasting relative transfection profiles are delineated.     

Preparative isolation of polyphenolic compounds from Vitis vinifera by centrifugal partition chromatography

This study deals with a centrifugal partition chromatography developed for the separation of phenolic compounds from Vitis vinifera. EtOAc grape seed extracts were separated using the solvent system hexane-ethyl acetate-ethanol-water (1:8:2:7; v/v) in two fractions: one containing about 75% of flavanol monomers (catechin and epicatechin) corresponding to 18% of crude extract and another fraction B-type dimers (22% of crude extract). From the stalk extracts, we could separate stilbenoid compounds (resveratrol and its oligomers; 12% of crude extract) which were eluted in less than 30 min from flavanols (which required a few hours of additional elution). Using the same solvent system but in different ratios (4:5:3:3; v/v), we isolated the trans-resveratrol (7@1000; 90% purity).     

Redox labeling of two antiepileptic drugs with metallocenes and their simultaneous detection by a Nafion-modified electrode

Two different cationic redox labels, i.e. a ferroceneammonium ion and a cobaltocenium ion, were covalently attached to two antiepileptics, phenytoin and phenobarbital, respectively. The two labeled drugs possess distinct standard redox potentials of 0.39 V for the phenytoin derivative and −0.92 V for phenobarbital derivative (vs Ag/AgCl, Cl⁻ 0.05 M ) at a carbon paste electrode. After preconcentration in a polyanionic Nafion-loaded carbon paste electrode the positively charged labeled phenytoin and phenobarbital derivatives could be simultaneously detected in concentration ranges which were relevant to the therapeutic ranges of the antiepileptics, with a view to a future dual-analyte immuno- assay. Square-wave voltammetry permitted detection limits of 5×10⁻⁸ M (for the phenytoin derivative) and 2.5×10⁻⁸ M (for the phenobarbital derivative) for non-simultaneous detection. © 1998 John Wiley & Sons, Ltd.     

Synthesis of vinyl pyrophosphonate analogues of farnesyl pyrophosphate: New potential inhibitors of farnesyl protein transferase

The synthesis of new bioisosteric analogues of farnesyl pyrophosphate where a vinyl pyrophosphonate replaced the pyrophosphate moiety is described. These compounds have been prepared using a Horner–Wadsworth–Emmons procedure and a modified Michelson reaction. They have been evaluated for the inhibition of farnesyl protein transferase. © 2002 Wiley Periodicals, Inc. Heteroatom Chem 13:654–661, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.10081     

Monitoring protein interactions in the living cell through the fluorescence decays of the cyan fluorescent protein.

Using fluorescence lifetime microspectroscopy and imaging techniques, we have studied the fluorescence of cyan fluorescent protein (CFP) transiently expressed in HEK-293 cells, in the presence or absence of its fluorescence resonance energy transfer (FRET) partner, yellow fluorescent protein (YFP). When the two proteins are attached through a 27-amino-acid linker, a 33 % average efficiency of intramolecular energy transfer is accurately determined inside the cell. Additionally, we observe a systematic quenching of the CFP fluorescence with increasing levels of protein expression. This quenching cannot be accounted for by formation of the previously described dimer of GFP-related proteins, since its magnitude is unchanged when the fluorescent proteins carry the mutation A206K shown to dissociate this dimer in vitro. Even when the intracellular protein concentration largely exceeds the in vitro dissociation constant of the dimer, self-association remains undetectable, either between free proteins or intramolecularly within the CFP-YFP construct. Instead, the detailed concentration effects are satisfactorily accounted for by a model of intermolecular, concentration-dependent energy transfer, arising from molecular proximity and crowding. In the case of CFP alone, we suggest that self-quenching could result from a pseudo-homo FRET mechanism between different, spectrally shifted emissive forms of the protein. These phenomena require careful consideration in intracellular FRET studies.     

Synthesis of P1 aspartate-based peptide acyloxymethyl and fluoromethyl ketones as inhibitors of interleukin-1β-converting enzyme

Improved procedures have been developed for the synthesis of P1 aspartate-based 2,6-dichlorobenzoyloxymethyl ketone 1 and fluoromethyl ketone 2, the prodrugs of two potent ICE-inhibitors. 1 was prepared from (R)-trans-4,5-O-isopropylidene-4,5-dihydroxy-2-pentenecarboxylic acid ethyl ester; 2 was obtained via a nitro-aldol condensation as key step from in situ generated fluoroacetaldehyde.