Rotational Correlation Time Tau Research Articles

The solvation of nitroxide radicals in ionic liquids studied by high-field EPR spectroscopy

Ionic liquids (ILs) feature a variety of properties that make them a unique class of solvents. To gain a better understanding of how ILs solvate compounds of different chemical structure, we used pulsed high-field electron paramagnetic resonance (EPR) spectroscopy at W-band (approximately 94 GHz) and continuous wave EPR at X-band (approximately 9.4 GHz) on three TEMPO-based spin probes with different substitutions at the 4-position: 4-R-2,2,6,6-tetramethylpiperidine-1-oxyl, with R = N(CH(3))(3)(+), Cat-1, R = COO(-), TEMPO-4-carboxylate, and R = OH, TEMPOL. The spin probes are dissolved in imidazolium based ILs with different alkyl chain lengths (-C(2)H(5), -C(4)H(9), -C(6)H(13)) and anions (BF(4)(-), PF(6)(-)) and also in molecular solvents (methanol, water-glycerol). X-Band EPR at RT shows that the reorientational motion of the charged spin probes in ILs is about fivefold slower than that of the TEMPOL. Moreover, anion variation from BF(4)(-) to PF(6)(-) in ILs most strongly slows down the rotational motion (as measured by the rotational correlation time tau(r)) of Cat-1, followed by TEMPOL, while tau(r) of TEMPO-4-carboxylate is least affected. The EPR parameters g(xx) and A(zz) (tensor elements of the g- and hyperfine tensor) are sensitive to environmental effects and are only fully resolved at the high field used in this study. Changes of g(xx) and A(zz) values of the Cat-1 in ILs and methanol are very small especially compared to that of TEMPO-4-carboxylate, indicating that Cat-1 is located in a polar region of the ILs resembling the situation in methanol. On the other hand, the g(xx) value of TEMPO-4-carboxylate is sensitive to the length of alkyl group which shows that TEMPO-4-carboxylate is close to the nonpolar region of ILs. The small differences in the chemical substitution of the spin probes used here are sufficient for the molecules to reside in different domains of different dielectric properties in ILs. Our combined results are in good agreement with a picture of a nanophase separation, in which the charged cations and anions form polar regions and the hydrophobic alkyl chains of the IL cations form non-polar regions.

Femtosecond polarization resolved spectroscopy: A tool for determination of the three-dimensional orientation of electronic transition dipole moments and identification of configurational isomers

A method is presented that combines femtosecond polarization resolved UV/visible pump-IR probe spectroscopy and density functional theory calculations in determining the three-dimensional orientation of an electronic transition dipole moment (tdm) within the molecular structure. The method is demonstrated on the approximately planar molecule coumarin 314 (C314) dissolved in acetonitrile, which can exist in two ground state configurations: the E- and the Z-isomer. Based on an exhaustive search analysis on polarization resolved measurement data for four different vibrational modes, it is concluded that C314 in acetonitrile is the E-isomer. The electronic tdm vector for the electronic S(0)-->S(1) transition is determined and the analysis shows that performing the procedure for four vibrational modes instead of the minimally required three reduces the 1sigma probability area from 2.34% to 2.24% of the solution space. Moreover, the fastest rotational correlation time tau(c) for the C314 E-isomer is determined to be 26+/-2 ps.

Dynamic spin label study of the barstar-barnase complex

The dynamic spin label method was used to study protein-protein interactions in the model complex of the enzyme barnase (Bn) with its inhibitor barstar. The C40A mutant of barstar (Bs) containing a single cysteine residue was modified with two different spin labels varying in length and structure of a flexible linker. Each spin label was selectively bound to the Cys82 residue, located near the Bn-Bs contact site. The formation of the stable protein complex between Bn and spin labeled Bs was accompanied by a substantial restriction of spin label mobility, indicated by remarkable changes in the registered EPR spectra. Order parameter, S, as an estimate of rapid reorientation of spin label relative to protein molecule, was sharply increasing approaching 1. However, the rotational correlation time tau for spin-labeled Bs and its complex with Bn in solution corresponded precisely to their molecular weights. These data indicate that both Bs and its complex with Bn are rigid protein entities. Spin labels attached to Bs in close proximity to an interface of interaction with Bn, regardless of its structure, undergo significant restriction of mobility by the environment of the contact site of the two proteins. The results show that this approach can be used to investigate fusion proteins containing Bn or Bs.

Unexpected Aggregation of Neutral, Xylene‐Cored Dinuclear GdIII Chelates in Aqueous Solution

We have synthesized ditopic ligands L(1), L(2), and L(3) that contain two DO3A(3-) metal-chelating units with a xylene core as a noncoordinating linker (DO3A(3-) = 1,4,7,10-tetraazacyclododecane-1,4,7-triacetate; L(1) = 1,4-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzene; L(2) = 1,3-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzene; L(3) = 3,5-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzoic acid). Aqueous solutions of the dinuclear Gd(III) complexes formed with the three ligands have been investigated in a variable-temperature, multiple-field (17)O NMR and (1)H relaxivity study. The (17)O longitudinal relaxation rates measured for the [Gd(2)L(1-3)(H2O)(2)] complexes show strong field dependence (2.35-9.4 T), which unambiguously proves the presence of slowly tumbling entities in solution. The proton relaxivities of the complexes, which are unexpectedly high for their molecular weight, and in particular the relaxivity peaks observed at 40-50 MHz also constitute experimental evidences of slow rotational motion. This was explained in terms of self-aggregation related to hydrophobic interactions, pi stacking between the aromatic linkers, or possible hydrogen bonding between the chelates. The longitudinal (17)O relaxation rates of the [Gd(2)L(1-3)(H2O)(2)] complexes have been analysed with the Lipari-Szabo approach, leading to local rotational correlation times tau(1)(298) of 150-250 ps and global rotational correlation times tau(g)(298) of 1.6-3.4 ns (c(Gd): 20-50 mM), where tau(1)(298) is attributed to local motions of the Gd segments, while tau(g)(298) describes the overall motion of the aggregates. The aggregates can be partially disrupted by phosphate addition; however, at high concentrations phosphate interferes in the first coordination sphere by replacing the coordinated water. In contrast to the parent [Gd(DO3A)(H2O)(1.9)], which presents a hydration equilibrium between mono- and dihydrated species, a hydration number of q = 1 was established for the [Ln(2)L(1-3)(H2O)(2)] chelates by (17)O chemical shift measurements on Ln = Gd and UV/Vis spectrophotometry for Ln = Eu. The exchange rate of the coordinated water is higher for [Gd(2)L(1-3)(H2O)(2)] complexes k(ex)(298) = 7.5-12.0 x 10(6) s(-1)) than for [Gd(DOTA)(H2O)](-). The proton relaxivity of the [Gd(2)L(1-3)(H2O)(2)] complexes strongly decreases with increasing pH. This is related to the deprotonation of the inner-sphere water, which has also been characterized by pH potentiometry. The protonation constants determined for this process are logK(OH) = 9.50 and 10.37 for [Gd(2)L(1)(H2O)(2)] and [Gd(2)L(3)(H2O)(2)], respectively.

Residue Specific Ribose and Nucleobase Dynamics of the cUUCGg RNA Tetraloop Motif by MNMR 13C Relaxation

The dynamics of the nucleobase and the ribose moieties in a 14-nt RNA cUUCGg hairpin-loop uniformly labeled with 13C and 15N were studied by 13C spin relaxation experiments. R1, R1rho and the 13C-[1H] steady-state NOE of C6 and C1' in pyrimidine and C8 and C1' in purine residues were obtained at 298 K. The relaxation data were analyzed by the model-free formalism to yield dynamic information on timescales of pico-, nano- and milli-seconds. An axially symmetric diffusion tensor with an overall rotational correlation time tau(c) of 2.31 +/- 0.13 ns and an axial ratio of 1.35 +/- 0.02 were determined. Both findings are in agreement with hydrodynamic calculations. For the nucleobase carbons, the validity of different reported 13C chemical shift anisotropy values (Stueber, D. and Grant, D. M., 2002 J. Am. Chem. Soc. 124, 10539-10551; Fiala et al., 2000 J. Biomol. NMR 16, 291-302; Sitkoff, D. and Case, D. A., 1998 Prog. NMR Spectroscopy 32, 165-190) is discussed. The resulting dynamics are in agreement with the structural features of the cUUCGg motif in that all residues are mostly rigid (0.82 < S2 < 0.96) in both the nucleobase and the ribose moiety except for the nucleobase of U7, which is protruding into solution (S2 = 0.76). In general, ribose mobility follows nucleobase dynamics, but is less pronounced. Nucleobase dynamics resulting from the analysis of 13C relaxation rates were found to be in agreement with 15N relaxation data derived dynamic information (Akke et al., 1997 RNA 3, 702-709).

Study of the miscibility and segmental motion of STMAA-PBMA polymer blends and semi-interpenetrating polymer networks by an ESR spin probe method

Linear polymer blends and semi-interpenetrating polymer networks (IPNs) with controlled hydrogen bonding interactions based on poly(styrene-co-methacrylic acid) (STMAA) and poly(butyl methacrylate) (PBMA) were studied by an ESR spin probe method. The observed composite ESR spectra with fast- and slow-motion components in all the samples were ascribed to two-phase morphology. For linear blends, the temperatures T(a) corresponding to appearance of the fast motion, T(d) corresponding to the disappearance of slow motion, T(5mT) and the rotational correlation times tau(c) increased with increasing carboxylic acid content in STMAA. It was concluded that the degree of mixing of the blends was improved with increasing carboxylic acid content, owing to the enhanced hydrogen bonding interactions between the carboxylic acids in STMAA and the ester groups in PBMA. With respect to semi-IPN samples, there existed a competition in the microphase structure between the intercomponent hydrogen bonding interactions, which improved the miscibility of the samples and the intracomponent cross-linking, which might lead to phase separation in the systems with strong specific interactions. When the semi-IPN contained 29 mol% carboxylic acid, the temperatures T(d), T(5mT) and tau(c) reached their minimum values, which indicated that the sample reached its maximum miscibility.

Determination of protein rotational correlation time from NMR relaxation data at various solvent viscosities

An accurate determination of the overall rotation of a protein plays a crucial role in the investigation of its internal motions by NMR. In the present work, an innovative approach to the determination of the protein rotational correlation time tau(R) from the heteronuclear relaxation data is proposed. The approach is based on a joint fit of relaxation data acquired at several viscosities of a protein solution. The method has been tested on computer simulated relaxation data as compared to the traditional tau(R) determination method from T(1)/T(2) ratio. The approach has been applied to ribonuclease barnase from Bacillus amyloliquefaciens dissolved in an aqueous solution and deuterated glycerol as a viscous component. The resulting rotational correlation time of 5.56 +/- 0.01 ns and other rotational diffusion tensor parameters are in good agreement with those determined from T(1)/T(2) ratio.

Relaxation, equilibrium oligomerization, and molecular symmetry of the VASP (336-380) EVH2 tetramer.

An investigation of the structural and dynamic properties of the C-terminal fragment of the human protein VASP (VASP 336-380) has been performed. Full length VASP has been shown to be tetrameric in solution, and the C-terminal 45 residues of the protein have been suggested to be responsible for the oligomerization. We have expressed and purified a C-terminal fragment of the human VASP protein from residue 336-380. It was found to form a stable domain in its own right. The fragment was shown by CD spectroscopy to form a helical structure, stable under a wide range of temperature and pH conditions. A (15)N-HSQC-experiment exhibits only one set of peaks, suggesting a high degree of symmetry for a putative oligomer. Measurements of the rotational correlation time tau(C) of the molecule and analytical ultracentrifugation data show VASP (336-380) to be entirely tetrameric in solution. The secondary structure was confirmed from a (15)N-NOESY-HSQC experiment and is completely alpha-helical. We conclude that VASP (336-380) forms a tetramer in solution via a coiled coil arrangement and is solely responsible for tetramerization of full-length VASP.

The impact of rigidity and water exchange on the relaxivity of a dendritic MRI contrast agent.

Variable-temperature, multiple magnetic field (17)O NMR, EPR and variable-temperature (1)H nuclear magnetic relaxation dispersion (NMRD) measurement techniques have been applied to Gadomer 17, a new dendritic contrast agent for magnetic resonance imaging. The macromolecule bears 24 Gd(dota)-monoamide chelates (dota=N,N',N",N"'-tetracarboxymethyl-1,4,7,10-tetraazacyclododecane) attached to a lysine-based dendrimer. (17)O NMR and (1)H NMRD data were analysed simultaneously by incorporating the Lipari-Szabó approach for the description of rotational dynamics. The water exchange rate k(298)(ex)was found to be (1.0 +/- 0.1) x 10(6) s(-1), a value similar to those measured for other Gd(dota)-monoamide complexes, and the activation parameters DeltaH++ =24.7 +/- 1.3 kJ mol(-1) and DeltaS++ = -47.4 +/- 0.2 JK(-1) mol(-1). The internal flexibility of the macromolecule is characterised by the Lipari-Szabó order parameter S(2)=0.5 and a local rotational correlation time tau(298)(l)= 760 ps, whereas the global rotational correlation time of the dendrimer is much longer, tau(298)(g)=3050 ps. The analysis of proton relaxivities reveals that, beside slow water exchange, internal flexibility is an important limiting factor for imaging magnetic fields. Electronic relaxation, though faster than in similar, but monomeric, Gd(III) chelates, does not limit proton relaxivity of this contrast agent (r(1)=16.5mM(-1)s(-1) at 298 K and 20 MHz). This analysis provides direct clues for the design of high-efficiency contrast agents.

Oligomerization of paramagnetic substrates result in signal amplification and can be used for MR imaging of molecular targets.

Magnetic resonance imaging (MRI) has evolved into a sophisticated, noninvasive imaging modality capable of high-resolution anatomical and functional characterization of transgenic animals. To expand the capabilities MRI, we have developed a novel MR signal amplification (MRamp) strategy based on enzyme-mediated polymerization of paramagnetic substrates into oligomers of higher magnetic relaxtivity. The substrates consist of chelated gadolinium covalently bound to phenols, which then serve as electron donors during enzymatic hydrogen peroxide reduction by peroxidase. The converted monomers undergo rapid condensation into paramagnetic oligomers leading to a threefold increase in atomic relaxtivity (R1/Gd). The observed relaxtivity changes are largely due to an increase in the rotational correlation time tau r of the lanthanide. Three applications of the developed system are demonstrated: (1) imaging of nanomolar amounts of an oxidoreductase (peroxidase); (2) detection of a model ligand using an enzyme-linked immunoadsorbent assay format; and (3) imaging of E-selectin on the surface of endothelial cells probed for with an anti-E-selectin-peroxidase conjugate. The development of "enzyme sensing" probes is expected to have utility for a number of applications including in vivo detection of specific molecular targets. One particular advantage of the MRamp technique is that the same paramagnetic substrate can be potentially used to identify different molecular targets by attaching enzymes to various antibodies or other target-seeking molecules.

Oligomerization of Paramagnetic Substrates Result in Signal Amplification and can be Used for MR Imaging of Molecular Targets

Magnetic resonance imaging (MRI) has evolved into a sophisticated, noninvasive imaging modality capable of high-resolution anatomical and functional characterization of transgenic animals. To expand the capabilities MRI, we have developed a novel MR signal amplification (MRamp) strategy based on enzyme-mediated polymerization of paramagnetic substrates into oligomers of higher magnetic relaxtivity. The substrates consist of chelated gadolinium covalently bound to phenols, which then serve as electron donors during enzymatic hydrogen peroxide reduction by peroxidase. The converted monomers undergo rapid condensation into paramagnetic oligomers leading to a threefold increase in atomic relaxtivity (R1/Gd). The observed relaxtivity changes are largely due to an increase in the rotational correlation time tau r of the lanthanide. Three applications of the developed system are demonstrated: (1) imaging of nanomolar amounts of an oxidoreductase (peroxidase); (2) detection of a model ligand using an enzyme-linked immunoadsorbent assay format; and (3) imaging of E-selectin on the surface of endothelial cells probed for with an anti-E-selectin-peroxidase conjugate. The development of "enzyme sensing" probes is expected to have utility for a number of applications including in vivo detection of specific molecular targets. One particular advantage of the MRamp technique is that the same paramagnetic substrate can be potentially used to identify different molecular targets by attaching enzymes to various antibodies or other target-seeking molecules.

Viscous flow and jump dynamics in molecular supercooled liquids. II. Rotations.

The rotational dynamics of a supercooled model liquid of rigid A-B dumbbells interacting via a Lennard-Jones potential is investigated along one single isobar. The time-temperature superposition principle, one key prediction of mode-coupling theory (MCT), was studied for the orientational correlation functions C(l). In agreement with previous studies we found that the scaling of C(l) in a narrow region at long times is better at high-l values. However, on a wider time interval the scaling works fairly better at low-l values. Consistently, we observed the remarkable temperature dependence of the rotational correlation time tau(1) as a power law in T-T(c) over more than three orders of magnitude and the increasing deviations from that law on increasing l (T(c) is the MCT critical temperature). For 0.7<T<2, good agreement with the diffusion model is found. For lower temperatures the agreement becomes poorer, and the results are also only partially accounted for by the jump-rotation model. The angular Van Hove function shows that in this region a meaningful fraction of the sample reorientates by jumps of about 180 degrees. The distribution of the waiting times in the angular sites cuts exponentially at long times. At lower temperatures it decays at short times as t(xi-1), with xi=0.34+/-0.04 at T=0.5, in analogy with the translational case. The breakdown of the Debye-Stokes-Einstein relation is observed at lower temperatures, where the rotational correlation times diverge more weakly than the viscosity.

Calix[4]arenes as selective extracting agents. An NMR dynamic and conformational investigation of the lanthanide(III) and thorium(IV) complexes.

The lanthanide and Th4+ complexes with calix[4]arene ligands substituted either on the narrow or at the wide rim by four coordinating groups behave totally differently as shown by an NMR investigation of the dia- and paramagnetic complexes. Solutions of complexes were prepared by reacting anhydrous metal perchlorate salts with the ligands in dry acetonitrile (CAUTION). Relaxation time T1 titrations of acetonitrile solutions of Gd3+ by calixarenes indicate that ligands subsituted on the narrow rim form stable 1:1 complexes whether they feature four amide groups (1) or four phosphine oxide functions. In contrast, a ligand substituted by four (carbamoylmethyl)-diphenylphosphine oxide moieties on the wide rim (3) and its derivatives from polymeric species even at a 1:1 ligand/metal concentration ratio. Nuclear magnetic relaxation dispersion (NMRD) curves (relaxation rates 1/T1 vs magnetic field strength) of Gd3+, Gd3+.1 and Gd3+.3 perchlorates in acetonitrile are analyzed by an extended version of the Solomon-Bloembergen-Morgan equations. A comparison of the calculated rotational correlation times tau r shows that ligand 3 forms oligomeric Gd3+ species. The chelates of ligand 1 are axially symmetric (C4 symmetry), and the paramagnetic shifts induced by the Yb3+ ion are accounted for quantitatively. The addition of water or of nitrate ions does not modify the geometry of the complex. The metal chelates of 3 and its derivatives adopt a C2 symmetry, and the paramagnetic shifts are interpreted on a semiquantitative basis only. Water and NO3- ions completely labilize the complexes of the heavy lanthanides. The very high selectivity of ligand 3 through the lanthanide series stems from a complex interplay of factors.

Backbone dynamics of the human CC-chemokine eotaxin.

Eotaxin is a CC chemokine with potent chemoattractant activity towards eosinophils. 15N NMR relaxation data have been used to characterize the backbone dynamics of recombinant human cotaxin. 15N longitudinal (R1) and transverse (R2) auto relaxation rates, heteronuclear ¿1H¿-15N steady-state NOEs, and transverse cross-relaxation rates (eta xy) were obtained at 30 degrees C for all resolved backbone secondary amide groups using 1H-detected two-dimensional NMR experiments. Ratios of transverse auto and cross relaxation rates were used to identify NH groups influenced by slow conformational rearrangement. Relaxation data were fit to the extended model free dynamics formalism, yielding parameters describing axially symmetric molecular rotational diffusion and the internal dynamics of each NH group. The molecular rotational correlation time (tau m) is 5.09 +/- 0.02 ns, indicating that eotaxin exists predominantly as a monomer under the conditions of the NMR study. The ratio of diffusion rates about unique and perpendicular axes (D parallel/D perpendicular) is 0.81 +/- 0.02. Residues with large amplitudes of subnanosecond motion are clustered in the N-terminal region (residues 1-19), the C-terminus (residues 68-73) and the loop connecting the first two beta-strands (residues 30-37). N-terminal flexibility appears to be conserved throughout the chemokine family and may have implications for the mechanism of chemokine receptor activation. Residues exhibiting significant dynamics on the microsecond-millisecond time scale are located close to the two conserved disulfide bonds, suggesting that these motions may be coupled to disulfide bond isomerization.

Internal motions of apo-neocarzinostatin as studied by 13C NMR methine relaxation at natural abundance.

Dynamics of the backbone and some side chains of apo-neocarzinostatin, a 10.7 kDa carrier protein, have been studied from 13C relaxation rates R1, R2 and steady-state 13C-(1H) NOEs, measured at natural abundance. Relaxation data were obtained for 79 nonoverlapping C alpha resonances and for 11 threonine C beta single resonances. Except for three C alpha relaxation rates, all data were analysed from a simple two-parameter spectral density function using the model-free approach of Lipari and Szabo. The corresponding C-H fragments exhibit fast (tau e < 40 ps) restricted libration motions (S2 = 0.73 to 0.95). Global examination of the microdynamical parameters S2 and tau e along the amino acid sequence gives no immediate correlation with structural elements. However, different trends for the three loops involved in the binding site are revealed. The beta-ribbon comprising residues 37 to 47 is spatially restricted, with relatively large tau e values in its hairpin region. The other beta-ribbon (residues 72 to 87) and the large disordered loop ranging between residues 97-107 experience small-amplitude motions on a much faster (picosecond) time scale. The two N-terminal residues, Ala1 and Ala2, and the C-terminal residue Asn113, exhibit an additional slow motion on a subnanosecond time scale (400-500 ps). Similarly, the relaxation data for eight threonine side-chain C beta must be interpreted in terms of a three-parameter spectral density function. They exhibit slower motions, on the nanosecond time scale (500-3000 ps). Three threonine (Thr65, Thr68, Thr81) side chains do not display a slow component, but an exchange contribution to the observed transverse relaxation rate R2 could no be excluded at these sites. The microdynamical parameter (S2, tau e and R2ex) or (S(slow)2, S(fast)2 and tauslow) were obtained from a straightforward solution of the equations describing the relaxation data. They were calculated assuming an overall isotropic rotational correlation time tau c for the protein of 5.7 ns, determined using standard procedures from R2/R1 ratios. However, it is shown that the product (1-S2) x tau e is nearly independent of tau c for residues not exhibiting slow motions on the nanosecond time scale. In addition, this parameter very closely follows the heteronuclear NOEs, which therefore could be good indices for local fast motions on the picosecond time scale.

Rotational and translational water diffusion in the hemoglobin hydration shell: dielectric and proton nuclear relaxation measurements

The dynamic properties of water in the hydration shell of hemoglobin have been studied by means of dielectric permittivity measurements and nuclear magnetic resonance spectroscopy. The temperature behavior of the complex permittivity of hemoglobin solutions has been measured at 3.02, 3.98, 8.59, and 10.80 GHz. At a temperature of 298 K the average rotational correlation time tau of water within a hydration shell of 0.5-nm thickness is determined from the activation parameters to be 68 +/- 10 ps, which is 8-fold the corresponding value of bulk water. Solvent proton magnetic relaxation induced by electron-nuclear dipole interaction between hemoglobin bound nitroxide spin labels and water protons is used to determine the translational diffusion coefficient D(T) of the hydration water. The temperature dependent relaxation behavior for Lamor frequencies between 3 and 90 MHz yields an average value D(298K) = (5 +/- 2) x 10(-10)m2 s-1, which is about one-fifth of the corresponding value of bulk water. The decrease of the water mobility in the hydration shell compared to the bulk is mainly due to an enhanced activation enthalpy.

Deuteron nuclear magnetic resonance study of the dynamic organization of phospholipid/cholesterol bilayer membranes: molecular properties and viscoelastic behavior

The influence of cholesterol on the dynamic organization of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayers was studied by deuteron nuclear magnetic resonance (2H NMR) using unoriented and macroscopically aligned samples. Analysis of the various temperature- and orientation-dependent experiments were performed using a comprehensive NMR model based on the stochastic Liouville equation. Computer simulations of the relaxation data obtained from phospholipids deuterated at the 6-, 13- and 14-position of the sn-2 chain and cholesterol labeled at the 3 alpha-position of the rigid steroid ring system allowed the unambiguous assignment of the various motional modes and types of molecular order present in the system. Above the phospholipid gel-to-liquid-crystalline phase transition, TM, 40 mol % cholesterol was found to significantly increase the orientational and conformational order of the phospholipid with substantially increased trans populations even at the terminal sn-2 acyl chain segments. Lowering the temperature continuously increases both inter- and intramolecular ordering, yet indicates less ordered chains than found for the pure phospholipid in its paracrystalline gel phase. Trans-gauche isomerization rates on all phospholipid alkyl chain segments are slowed down by incorporated cholesterol to values characteristic of gel-state lipid. However, intermolecular dynamics remain fast on the NMR time scale up to 30 K below TM, with rotational correlation times tau R parallel for DMPC ranging from 10 to 100 ns and an activation energy of ER = 35 kJ/mol. Below 273 K a continuous noncooperative condensation of both phospholipid and cholesterol is observed in the mixed membranes, and at about 253 K only a motionally restricted component is left, exhibiting slow fluctuations with correlation times of tau R perpendicular greater than 1 microsecond. In the high-temperature region (T greater than TM), order director fluctuations are found to constitute the dominant transverse relaxation process. Analysis of these collective lipid motions provides the viscoelastic parameters of the membranes. The results (T = 318 K) show that cholesterol significantly reduces the density of the cooperative motions by increasing the average elastic constant of the membrane from K = 1 x 10(-11) N for the pure phospholipid bilayers to K = 3.5 x 10(-11) N for the mixed system.

Molecular motion of drugs in hydrocolloids measured by electron paramagnetic resonance.

The effects of a polymer, the Li-salt copolymer of methylmethacrylic acid, and its methyl ester on the motion of drug molecules in hydrocolloids were studied. The investigation was carried out by means of electron paramagnetic resonance (EPR) using the model nitroxide tempol, and the spin-labeled drugs lidocaine (sl-lid) and dexamethasone (sl-dex). Synthesis of sl-dex was performed. Spin-labeled molecules dissolved in hydrocolloids undergo a fast reorientation motion. The decreasing order of rotational correlation times (tau)--sl-dex greater than sl-lid greater than tempol--suggests that the size and the shape of the molecules strongly affect their motion. The inhibition of motion of larger molecules depends also on their flexibility. The tau values indicate proportionality of the microviscosity of hydrocolloids to the polymer concentration. Rotational motion is dependent on the local environment conditioned by the free spaces between polymer molecules.

Study of the Mode of Action of Some Nitrodiphenyl Ethers

Nitrosoderivatives of the nitrodiphenyl ether herbicides (nitrofen, bifenox) have been studied. UV irradiation in different organic solvents gives degradation products. In buffered aqueous media, in the presence of chloroplasts and spin traps such as DMPO, hydroxy and peroxy radicals have been characterized. In organic media and in the presence of spin traps such as DMPO, PBN, 4-POBN, solvent radicals (.CHCl2, .CCl3, .CH2 [symbol: see text]) have been formed. Nitro-derivatives have been studied under UV irradiation and in the presence of tetramethylethylene (TME), alkenylhydroxylamines are formed which autoxidize in nitroxide radicals. The formation of the stable nitroxide radical occurs in the dark process after continuous irradiation. The intensity of the signal decreases strongly when a new irradiation is applied. Radical species, with analogous ESR spectral characteristics are formed on reaction with nitrodiphenyl ethers and fatty acids. The reactivity of these herbicides in micellar media (SDS, Brij 35, and CTAB) has been investigated. The kinetics of formation of the ESR signal corresponding to the photoreduction of the nitrodiphenyl ether in the presence of TME behave differently in a micellar environment as compared to solution. The intensity of the formation of the nitroxide increases under irradiation and decreases in the dark; the rotational correlation time tau c has been determined for each type of micelle. Synthetic nitrosodiphenyl ether made by the reduction of nitrodiphenyl ether using hydrogen gas and PtO2 as a catalyst gives the corresponding amine, which is oxidized with meta-chloroperbenzoic acid (m.CPBA). The nitrosodiphenyl ether in the presence of soja azolectin liposome containing a fluorescent probe has been analysed.(ABSTRACT TRUNCATED AT 250 WORDS)

Temperature-dependent molecular motions and phase behavior of cholesteryl ester analogues.

The phase behavior and temperature-dependent molecular motions of three cholesteryl ethers (caproyl, myristyl, oleyl) and a cholesteryl carbonate (oleyl) were characterized. The properties of each ether were qualitatively similar to, but quantitatively different from, those of the corresponding cholesteryl ester. For example, cholesteryl oleyl ether exhibited the same phase transitions as cholesteryl oleate, but at much lower temperatures (e.g., the ether isotropic liquid to cholesteric transition is at 29 degrees C). 13C NMR spectra of ethers in the isotropic liquid and liquid crystalline phases were similar to those of the ester analogue. However, near the liquid to liquid crystalline transition, the steroid ring C3 and C6 linewidths, the C3/C6 linewidth ratio, and the steroid ring rotational correlation times tau rx and tau rz calculated from the linewidths were larger for the ether than the ester analogue. The oleyl carbonate had qualitatively different properties from its analogues (e.g., stable vs. metastable cholesteric and smectic phases). Quantitative results (e.g., relatively long tau rx and tau rz in the isotropic liquid phase) for the carbonate were also distinct from those of both the ester and ether analogues. A comparison of analogues in which the polar linkage is the only structural variable yielded insights into the intermolecular interactions which influence phase behavior.