Spin probing hydrophobic probes

In this section, the EPR spectroscopic characterization of thermoresponsive polymeric systems is presented. The polymeric systems are water-swollen at lower temperatures and upon temperature increase the incorporated water is driven out and the system undergoes a reversible phase separation. Simple CW EPR spectroscopy (see above), carried out on a low-cost, easy-to-use benchtop spectrometer, is used here to reveal and characterize inhomogeneities on a scale of several nanometers during the thermal collapse. Further, neither any physical model of analysis nor chemical synthesis to introduce radicals had to be utilized. Adding amphiphilic TEMPO spin probes as guest molecules to the polymeric systems leads to self-assembly of these tracer molecules in hydrophilic and hydrophobic regions of the systems. These probes in different environments can be discerned and one... 

The spectral parameters for component A again coincide with those of TEMPO in pure water ( a 48.3 MHz), i.e., this spin probe is located in a strongly hydrated, hydrophilic environment. The observed decrease of ai o by 3.7 MHz for species B ( hnal at 65 °C) is indicative of much more hydrophobic and less hydrated surroundings for these spin probes (comparable to chloroform or tert-hutyl alcohol [83]). At temperatures below the collapse temperature Tq, only the hydrophilic spectral component A is observed since all dendritic units are water-swollen. Above the critical temperature of 33 °C an increasing fraction of hydrophobic species B is observed with increasing temperature. The dehydration of the dendritic units thus leads to a local phase separation with the formation of hydrophobic cavities. Unlike in PNIPAAM hydrogels (Sect. 3.1), here hydrophobic regions are not observed below the macroscopic collapse temperature. 

Fig. 8 Model of the collapse of thermoresponsive dendronized polymers as seen by EPR spin probes few individual hydrophobic and dynamic patches of 5 nm are sufficient to achieve a macroscopic collapse at the cloud point. Only at temperatures well above a static state (on EPR time scales) is reached...

Ottaviani and various coworkers have in the past often used a spin probing/ spin labeling approach to study self-assembled soft matter systems. In a recent paper they describe the complexes formed between cationic surfactants and hydrophobic ally modified anionic polymers. Analyzing the changes in dynamics as well as environment (hydrophobicity) of the long-chain fatty acid spin probe... 

Similarly, Wasserman and coworkers have studied a wide selection of polymeric materials in aqueous solution that are associative of some kind, i.e., that form some sort of self-assembly through non-covalent interactions [96]. Their study mainly deals with hydrogels of hydrophobically modified polymers, aqueous solutions of polymeric micelles created by block copolymers, and hydrogels based on poly (acrylic acid) and macrodiisocyanates. The spin probes of choice were hydrophobic, such as 5- and 16-DSA (see Eig. 2) or even spin labeled polymers. It was, e.g., possible to screen for the effect of chemical stmcture on the gel formation by recording and understanding the local mobility of the hydrophobic, long chain spin probes as a function of temperature. 

Lowe, T. L., Virtanen, J. and Tenhu, H., Interactions of drugs and spin probes with hydrophobically modified polyelectrolyte hydrogels based on N-isopropylacrylamide. Polymer, 40, 2595-2603 (1999). 

Therefore, a purpose of the present work was to study the effect of DPhO and BM-DPhO in a wide range of concentration (10 -10 mol/1) on the endoplasmic reticulum membranes (microsomes) isolated from Balb-line mice. Electron paramagnetic resonance (EPR) technique and spin-probe method were used to study the dynamic structure of deep hydrophobic and surface lipid regions of microsomal membranes. We suggested the different effects of DPhO and IM-DPhO on the membrane lipids structure, because iod-methylate derivative is charged. 

FIGURE 1 Structural formula of spin probes 5-DSA (left side) and 16-DSA (right side), EPR spectra in surface and deep hydrophobic regions of lipid bylaer of itiicrosonial membranes correspondingly. 

The spin probes 5- and 16-DSA are localized in the different regions of membrane lipids 5-DSA in surface lipids at depth of 8A° and hydrophobic nitroxyl radical 16-DSA penetrates in lipid bilayer to the depth more than 20A° [14, 16], A motion of 16-DSA can be characterized by rotation correlation time (x) by which a microviscosity value is estimated and anisotropic rotation of 5-DSA by order parameter (S) [15,16],... 

By spin probes 5- and 16-DSA it was found that DPhO and its iod-methylate derivate (IM-DPhO) increased the microviscosity (t) in the depth hydrophobic regions and rigidity (S) of surface membrane lipids depending on the concentration by nonlinear manner. 

In order to avoid any possible perturbance caused by a hydrophobic chain of the probe. Ristori examined the state of water in the interlamellar regions of perfluoropolyether ammonium carboxylates by using the corresponding Cu(II) [159a] and Mn(II) [159b] salts as the spin probe. 

We would like to stress that in all these publications the authors investigated peculiarities of the rotational and translational diffusion of spin-probe molecules in various room temperature ionic liquids, comp)ared them with molecular dynamics in common organic solvents. Correlations with Stokes-Debye-Einstein or Stokes-Enstein laws were foimd. Areas in RTILs (polar, non-polar), in which spin probes (hydropElic, charged, hydrophobic) are localized were determined. Just recently, attention of the scientists was attracted to another type of molecular motions in the ionic liquids (Tran et al., 2007a, 2009). Such processes as well as solvent effects on them can be examined in detail by EPR sp>ectroscopy with the use of stable nitroxide biradicals (Parmon et al., 1977a, 1980). 

CONTENTS Preface, C. Allen Bush. Thermodynamic Solvent Isotope Effects and Molecular Hydrophobicity, Terrence G. Oas and Eric J. Toone. Membrane Interactions of Hemolytic and Antibacterial Peptides, Karl Lohner and Richard M. Epand. Spin-Labeled Metabolite Analogs as Probes of Enzyme Structure, Chakravarthy Narasimhan and Henry M. Miziorko. Current Perspectives on the Mechanism of Catalysis by the Enzyme Enolase, John M. Brewer and Lukasz Leb-ioda. Protein-DNA Interactions The Papillomavirus E2 Proteins as a Model System, Rashmi S. Hedge. NMR-Based Structure Determination for Unlabeled RNA and DNA, Philip N. Borer, Lucia Pappalardo, Deborah J. Kenwood, and Istvan Pelczer. Evolution of Mononuclear to Binuclear CuA An EPR Study, William E. Antholine. Index. 

Franklin JC, Cafiso DS, Flewelling RF, et al. Probes of membrane electrostatics synthesis and voltage-dependent partitioning of negative hydrophobic ion spin labels in lipid vesicles. Biophys J 1993 64(3) 642-653. 

To determine the shape of the hydrophobic barrier of bilayer membranes, fatty acids and PC molecules spin labeled with nitroxides at various positions along the lipid chains were diffused into vesicles and their solvent-sensitive isotropic coupling constants were measured [54]. Results are plotted in Figure 5 in terms of distance of the probe from the bilayer center. Also shown is the profile of the dielectric constant along the membrane normal evaluated from the fluorescence lifetime distribution of fluorescence probes in PC liposomes [55]. These data correlate well with results from neutron diffraction studies that map the positional distribution of water and lipid moieties along the bilayer normal [56]. 

---