Probing the mechanism of transport and compartmentalisation of polyamines in mammalian cells.

BACKGROUND: Many mammalian cells possess an active polyamine uptake system but little is known about the molecular mechanism of this transporter. The fate of polyamines taken up from the medium and the relationship to polyamine homeostasis remains to be fully established. The aim of this study was to develop a range of modified polyamines, particularly ligands incorporating a fluorophore, to explore the structural tolerances of the polyamine transport system and to probe the intracellular location of polyamines acquired from the medium. RESULTS: We synthesised a wide range of polyamine analogues incorporating cytotoxic agents, fluorescent chromophores and bulky substituents. All of these analogues have been shown to be good competitive inhibitors of spermidine uptake in a range of mammalian cells. Direct evidence for uptake of the fluorescent polyamine analogues and their subcellular distribution was obtained from confocal laser scanning fluorescence microscopy, which showed that they accumulated in granular structures within the cytoplasm and not in the nucleus. We demonstrated that their uptake is through the polyamine transport system by showing that pretreatment with DFMO, a potent inhibitor of polyamine biosynthesis, led to enhanced uptake, and cells deficient in the polyamine transport system did not accumulate these polyamine analogues. CONCLUSIONS: The polyamine transport system has a surprisingly broad structural tolerance. Fluorophore-containing polyamine analogues derived from the extracellular pool are located in granular structures within the cytoplasm and not to any great extent in the nuclei of mammalian cells. These observations might be consistent with a mechanism involving receptor-mediated endocytosis, and the granular 'structures' seen might reflect polyamine compartmentalisation within vesicles.     

Self-chemical ionization of diethylzinc

The self-chemical ionization of diethylzinc is examined by Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry and semiempirical molecular orbital calculations. Electron impact of diethylzinc neutral produces the radical cation, C(4)H(15)Zn(+) (m/z x 122), which reacts further with the neutral (C(2)H(5))(2)Zn to give the following product ions: Zn(+) (m/z x 64), C(2)H(5)Zn(+) (m/z x 93), C(4)H(9)Zn(+) (m/z x 121), C(4)H(11)Zn(2)(+) (m/z x 187), and C(6)H(15)Zn(2)(+) (m/z x 215). To determine the structure and pathways for production of these ions, monoisotopic (12)C(4)H(15)(64)Zn(+), (64)Zn(+) and (12)C(2)H(5)(64)Zn(+) were individually isolated and reacted with the neutral background. We also performed semiempirical molecular orbital calculations (ZINDO/1). The molecular orbital calculations and experimental data are consistent in predicting that the ethyl group on the diethylzinc cation carries the positive charge. Copyright 1999 John Wiley & Sons, Ltd.     

Effects of added dotab on thecmc and enthalpy of micelle formation at 298.2 K for CTAB(aq)

In a titration calorimetric study an aqueous solution held in a syringe and containing hexadecyltrimethylammonium bromide (CTAB; 15.4×10^–3 mol dm^–3) is injected in aliquots (5×l0^–3 dm³) into a sample cell containing initially water. Analysis of the data shows that thecmc equals 0.97×l0^–3 dm^–3 and the enthalpy of micelle formation equals –10.3 kJ mol^–1. When the solution in the syringe is replaced by a mixed surfactant solution, CTAB+dodecyltrimethylammonium bromide, at the same total concentration of surfactant, thecmc of CTAB decreases gradually with increasing mole fraction of DOTAB but the enthalpy of CTAB micelle formation is hardly affected. We conclude, therefore, that incorporation of DOTAB monomers into the CTAB micelles stabilizes entropically the CTAB micelles.We thank EPSRC for their support; the Commonwealth Scholarship Commission for an award to MCSS and the Royal Society for a grant awarded to PMC for the purchase of the Titration Microcalorimeter.     

Differential scanning and titration calorimetric studies of macromolecules in aqueous solution

An enormous amount of detailed information can be obtained concerning macromolecules in aqueous solution using data obtained from differential scanning and titration microcalorimetry. This claim is supported by reference to examples taken from recent work concerned with micelles (e.g. CTAB), vesicles (e.g. DOAB and DDP) and enzymes (e.g. CAT and DNA gyrase).     

Combinations of titanium(IV) oxide,iron(III) oxide and molybdenum(VI) oxide as flame retardants and smoke suppressants for thermoplastic polymers

An inert metal oxide (TiO₂), a catalytic oxide (Fe₂O₃) and an oxide which forms volatile halides (MoO₃) have been incorporated into polypropylene and polystyrene on their own, in combination with one another, and in the absence and presence of a halogen compound (Cereclor 70 or decabromobiphenyl oxide). Studies of the resulting flame-retardant and smoke-suppressant effects of the additive systems have been carried out by means of triangular diagrams. None of the systems investigated has any sizeable effect on the flammability of polypropylene, the limiting oxygen indices being raised by no more than 8 units. A positive interaction between Cereclor 70 and molybdenum oxide results however both in an improvement in the degree of flame retardance of this polymer and in a decrease in its already rather low smoke-producing tendency. These additive systems have proved more effective in the case of polystyrene. The decreased flammability manifests itself as an increase of up to 12 LOI units, while the maximum smoke density resulting from the combustion of the polymer is considerably decreased. In some cases the maximum smoke density, D_s, is less than 5% of the value for the pure polymer. The most efficient flame-retardant and smoke-suppressant systems generally involve molybdenum oxide and these are, in particular, ternary systems containing also another metal oxide and decabromobiphenyl oxide. Iron(III) oxide has little effect on its own but nevertheless considerably enhances the activity of the other metal oxides.     

Metal chelates as flame retardants and smoke suppressants for thermoplastic polymers

Metal chelates have been incorporated into polystyrene and polypropylene to study their effects on the combustion of these polymers, in the absence and presence of a chlorinated hydrocarbon. In particular, the effect of ferrocene on the flammability of the polypropylene-Cereclor 70 system has been studied comprehensively by a triangular diagram method. The additives have been shown to interact synergistically to increase char and thus produce a system of low flammability; the Cl/Fe ratio needed for optimum flame retardance is considerably higher than stoichiometric. It has been found that, if the presence of the chlorine compound decreases the amount of smoke produced from a polymer-metal chelate system, this is indicative of some degree of flame-retardant activity of the metal-halogen system.     

Binary mixtures of metal compounds as flame retardants for organic polymers

Studies have been made of the effect on the flammability of thermoplastic polymers of the partial or total replacement of one metal compound by another in the presence also of a suitable halogen compound; particular attention has been paid to systems where the primary flame retardant is antimony(III) oxide. With each binary metal compound system investigated, ten different compositions have been chosen so as to provide a symmetrical arrangement of points within a triangular design; resulting calculated values of the limiting oxygen index for each polymer-flame retardant system for a given polymer are shown as a graphical contour analysis. Comprehensive studies of several systems show that both iron(III) oxide and aluminium oxide monohydrate can significantly enhance the flame-retardant action of antimony(III) oxide but that several other metal compounds, although not as effective as Sb₂O₃, may nevertheless be used as adequate partial replacements for it. The Fe₂O₃-SnO₂-H₂O system can also act as an effective flame retardant under certain conditions. The SnOZnO system perhaps best illustrates the importance of the polymer substrate and of the total additive loading as factors controlling the flame-retardant effectiveness. For all the systems studied, however, ABS is a much better substrate than HDPE. The results of a reasonably detailed study of the flame retardance conferred by several different compositions of a binary metal compound mixture give a much more reliable indication of the effects on polymer flammability of the constituent metal compounds than are obtained simply by replacement of a given concentration of one compound by another.     

Paper chromatography of fatty acids and halogenated organic acids

A paper chromatographic method is described for the identification of the volatile fatty acids and some of their bromo- and iodo-derivatives. Some new spraying reagents are suggested which enable the different classes of acids to be detected in presence of one another.     

Kinetics of wetting of a compacted powder by aqueous solutions of surface active agents

Summary The capillary rise of aqueous solutions of anionic wetting agents into a compacted powder of an organic chromium complex is discussed on the basis of the Washburn-Rideal equation expanded to account for the porous structure parameters.At the porosity 1 —/6, corresponding to the loosest packing of monodisperse spheres the penetration rate is found to be mostly governed by the effective tensions operative at the moving three phase line of contact in case no deflocculation intervenes.

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Zusammenfassung Der Kapillaraufstieg wässeriger Lösungen anionischer Netzmittel in gepreßtem Pulver eines organischen Chrom-Komplexes wird durch die erweiterte Gleichung vonWashburn-Rideal wiedergegeben, in der die Parameter der porösen Struktur mit einbezogen werden.Bei der Porosität 1 —/6, die der lockersten Anordnung gleichförmiger Kugeln entspricht, wird die Eindringgeschwindigkeit meistens durch die an der sich bewegenden Dreiphasengrenze wirkenden Grenzflächenspannungen bestimmt, wenn keine Peptisation eintritt.

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Paper presented at the XII Intern. Congr. Fat Research, Milan, September 1974.