High Counterion Concentration Research Articles

Fabrication and Characterization of a Reversed-Phase/Strong Cation Exchange Stationary Phase Gradient

Continuous stationary phase gradients for liquid chromatography (LC) have been recently shown to be a promising method of altering selectivity. In this work, we present the first multicomponent continuous stationary phase gradient for separations involving both reversed-phase (RP) and strong cation exchange (SCX) mechanisms. These columns are fabricated using a two-step methodology based on controlled rate infusion (CRI). First, destructive CRI creates a gradient of excess silanol groups along a uniform C18 column. Next, these columns are infused with 3-mercaptopropyltrimethoxysilane (MPTMOS), which bonds to the excess silanol groups. The terminal thiols of the MPTMOS ligands are oxidized with H2O2 to create the sulfonate functionality (SO3−) needed for SCX separations. The success of the modification procedure is characterized by thermogravimetric analysis and X-ray photoelectron spectroscopy. The stability of the C18-SO3− gradients were found to have less than 5 % retention loss and the column-to-column reproducibility had a relative standard deviation under 9 %. The peak asymmetry factors for seven biogenic amines were found to be between 1.03 ± 0.04 to 1.30 ± 0.02, which illustrates minimal peak tailing due to poor packing and residual silanol groups. Characterization of the gradient columns using an isocratic mobile phase showed a unique elution order compared to a uniform C18 and SO3− columns. At lower counterion concentrations, more than 48 % of the overall retention on the gradient stationary phase is due to a SCX mechanism. Meanwhile, the RP mechanism was shown to predominate at higher counterion concentrations on the gradient columns (SCX retention contribution less than 40 %). Coupling the stationary phase gradient to a salt gradient in the mobile phase showed that the gradient phase provides a unique, intermediate selectivity to the uniform C18 and SO3− columns. Under an ACN mobile phase gradient, a significant increase (p < 0.003) in the retention times of three biogenic amines (15 – 16 %) was observed when the multicomponent gradient was oriented to have a high SO3− ligand density near the detector. This work serves as a proof-of-concept design for a multicomponent stationary phase gradient to continue fundamental studies into retention and encourage novel applications.

Unusual stability exhibited by (AT)XN12(AT)Y motif associated with high fetal hemoglobin levels

Quasi-palindromic sequences (AT)XN12(AT)Y present in HS2 (hypersensitive site 2) of the human β-globin locus are known to be significantly associated with increased fetal hemoglobin (HbF) levels. High HbF levels in some adults arise due to pathological conditions such as sickle cell disease and β-thalassemia. However, elevated levels of HbF are also associated with a reducing morbidity and mortality in patients with β-thalassemia and thus ameliorate the severity of the disease. Using gel-electrophoresis, ultraviolet (UV)-thermal denaturation, and circular dichroism (CD) techniques, we demonstrated that it exhibits a hairpin-duplex equilibrium. Intramolecular species (hairpin) were observed in both low and high salt concentrations in gel assay studies displaying the unusual stability of intramolecular species even at the high counter-ion concentration. The unusual stability of hairpin secondary structures was also demonstrated by the monophasic nature of the melting profiles for the oligonucleotides which persisted at low as well as high salt and oligomer concentrations. Change in CD spectra as a function of oligomer concentration indicates that the bimolecular duplex formation is selectively favored over monomolecular hairpin formation at and above 9 µM oligomer concentration. Thus, we hypothesize that imperfect inverted repeat sequence (AT)XN12(AT)Y of HS2 of β-globin gene LCR forms the unusually stable hairpins which may result in the formation of a cruciform structure that may be recruited for binding by various nuclear proteins that could result in elevated HbF levels.Communicated by Ramaswamy H. Sarma.

Effect of salt counterions on the physicochemical characteristics of novel green surfactant, ethane-1,2-diyl bis(N,N-dimethyl-N-tetradecylammoniumacetoxy) dichloride

Abstract In this research article we have examined the effect of chloride (Cl−), sulphate (SO42−), phosphate (PO43−), anthranilate (An−) and tosylate (Tos−) counterions on the physicochemical characteristics of green gemini surfactant, ethane-1,2-diyl bis(N,N-dimethyl-N-tetradecylammoniumacetoxy) dichloride (14-E2-14) through a multidimensional approach. Substantial changes were observed in the physicochemical parameters. Counterions were found to cause reduction in critical micellar concentration (CMC) and minimum area per head group ( A min ) of the 14-E2-14 gemini surfactant. However, increase in aggregation number (Ng) and surface excess concentration ( Γ max ) of the 14-E2-14 surfactant was observed upon increasing the counterion concentrations. The Gibbs free energies of micellization (ΔG°mic) and adsorption (ΔG°ads) were found to be negative, indicating that 14-E2-14 micellization is spontaneous. The order of efficacy of counterions to influence the micellar characteristics was: An− > Tos− > PO43− > SO42− > Cl−. Further, 1H NMR results demonstrated morphological transitions (sphere-to-rod) and newer phase formation. The UV–vis and FT-IR results delineate intercalation of organic counterions (An− and Tos−) into the palisade layer of 14-E2-14 micelles. TEM micrographs show emergence of non-spheroidal micelles at higher counterion concentrations. This study could be significant to surfactant science scientists in tailoring green surfactants with desirable physicochemical characteristics, so that special-characteristic-tailored surfactants could be exploited in designing and compilation of environmentally benign industrial, cosmetic and biomedical products.

The effect of pH on charge inversion and condensation of DNA.

Charge inversion and condensation of DNA in solutions of trivalent and quadrivalent counterions are significantly influenced by the pH value of the solution. We systematically investigated the condensation and charge compensation of DNA by spermidine, hexammine cobalt(iii) (cohex, [Co(NH3)6](3+)) and spermine in solutions of a wide range of pH values from 3 to 9.3 by dynamic light scattering, magnetic tweezers, and atomic force microscopy. In trivalent counterion solution, we found that there is a critical concentration (0.75 mM for cohex and 0.5 mM for spermidine), under which the electrophoresis mobility of DNA initially increases, reaches a maximum, and finally decreases when the pH value is decreased. In contrast, above the critical concentration, the electrophoretic mobility of DNA increases monotonously with decreasing pH value of the solution. The corresponding condensing force has the same dependence on the pH value. However, for the case of quadrivalent counterions, the electrophoretic mobility of DNA is monotonously promoted by lowering the pH value of the solution at any concentration of counterions in which charge inversion of DNA may occur. In atomic force microscopy images and force spectroscopy of magnetic tweezers, we found that maximal charge neutralization and condensation force correspond to the most compact DNA condensation. We propose a mechanism of promoting DNA charge neutralization: small and highly mobile hydrogen ions tend to attach to the DNA-counterion complex to further neutralize its remaining charge, which is related to the surface area of the complex. Therefore, this further neutralization is prominent when the complex is toroidal which corresponds to the case of mild ion concentration while it is less prominent for more compact globules or rod complexes at high counterion concentration.

Synthesis of Platinum Catalysts over Thick Slurries of Oxide Supports by Strong Electrostatic Adsorption

AbstractPlatinum uptake by strong electrostatic adsorption on alumina, silica, and titania supports was studied at different slurry thicknesses (or surface loadings). For the anionic platinum chloride precursor, a high counterion concentration at high surface loadings hindered adsorption on the high point of zero charge alumina support, as predicted by the revised physical adsorption model. Owing to this adsorption retardation in thicker slurries, a highly dispersed Pt/Al2O3 catalyst could only be prepared with a lower weight fraction of Pt than that in thin slurries. However, for cationic platinum tetraamine complexes, the effect of counterions in retarding the adsorption at high surface loadings was unexpectedly diminished. As a result, it was possible to synthesize highly dispersed Pt/SiO2 even at a maximum slurry thickness (incipient wetness) without having to sacrifice the Pt loading.

Adsorption of an anionic surfactant at air-liquid and different solid-liquid interfaces from solutions containing high counter-ion concentration

The self-assembly (adsorption) of the anionic surfactant, sodium dodecylbenzenesulphonate (SDBS), at air-liquid and different (octadecanethiol, β-mercaptoethanol, α-lipoic acid) solid-liquid interfaces from aqueous solutions containing high concentrations of counter-ion has been investigated. SDBS adsorption at the solid-liquid interfaces was obtained using surface plasmon resonance (SPR) while its adsorption at the air-liquid interface was extracted from surface tension measurements. The results have demonstrated that SDBS packing at the air-liquid interface is similar to its packing at the hydrophobic octadecanethiol-liquid interface. Additionally, SDBS packing at the three solid-liquid interfaces increases with increasing surface hydrophobicity, irrespective whether the surface is neutral or negatively charged. Nonetheless, SDBS adsorption is always within monolayer coverage (no evidence of bilayer or admicelle formation). The results have also revealed that SDBS affinity for the solid-liquid interfaces increases with increasing surface hydrophobicity. Furthermore, SDBS affinity for the air-liquid interface is more than 10-fold its affinity for the solid-liquid interfaces.

Counterion and Polythymidine Loop-Length-Dependent Folding and Thermodynamic Stability of DNA Hairpins Reveal the Unusual Counterion-Dependent Stability of Tetraloop Hairpins

Stem-loop DNA hairpins containing a 5-base-pair (bp) stem and single-stranded polythymidine loop were investigated using thermodynamic melting analysis and stopped-flow kinetics. These studies revealed the thermodynamic stability and folding kinetics as a function of loop length and counterion concentration. Our results show the unusually high thermodynamic stability for tetraloop or 4 poly(dT) loop hairpin as compared with longer loop length hairpins. Furthermore, this exceptional stability is highly counterion-dependent. For example, in the higher counterion concentration regime of 50 mM NaCl and above, the tetraloop hairpin displays enhanced stability as compared with longer loop length hairpins. However, at lower counterion concentration of 25 mM NaCl and below, the thermal stability of tetraloop hairpin is consistent with the longer loop hairpins. The enhanced stability of tetraloop hairpins at higher counterion concentration can be explained on the basis of the combined entropic effect of loop closure as well as base stacking in the loop regions. The stability of longer loop length hairpins at all counterion concentrations as well as tetraloop hairpin at lower counterion concentration can be explained on the basis of entropic effect of loop closure alone. The thermodynamic parameters at lower and higher counterion concentrations were determined to quantify the enhanced stability of base-stacking effects occurring at higher counterion concentrations. For example, for 100 mM NaCl, excess Gibbs energy and enthalpy due to base stacking within the tetraloops were measured to be -1.2 ± 0.14 and -3.28 ± 0.32 kcal/mol, respectively, whereas, no excess of Gibbs energy and enthalpy was observed for 0, 5, 10, and 25 mM NaCl. These findings suggest significant base-stacking interactions occurring in the loop region of the tetraloop hairpins at higher counterion concentration and less significant base-stacking interactions in the lower counterion concentration regime. We suggest that at higher counterion concentrations, hydrophobic collapse of the nucleotides in the loop may be enhanced due to the increased polarity of the solvent, thereby enhancing base-stacking interactions that contribute to unusually high stability.

Thermodynamically unfavorable DNA hybridizations can be made to occur by a water to ice phase change

In an apparent contradiction to Debye–Hückel theory, it was possible to hybridize DNA in solutions of Milli-Q water (resistivity>18MΩcm−1) containing no added ions. This was demonstrated by hybridizing four bi-complementary DNA sequences to form an ‘X’ shape, as indicated by acrylamide gel electrophoresis. The requirement for hybridization was that a water-to-ice phase change should occur. Comparative experiments, using freezing by liquid nitrogen and thawing at different temperatures, showed that hybridization could take place during either the freezing or thawing process provided either was slow enough. We speculate that the low solubility of DNA in ice creates liquid inclusions of extremely high DNA and counter-ion concentration prior to complete freezing, and that hence in these inclusions hybridization was actually in accordance with Debye–Hückel theory.

The Impact of Multivalent Counterions, Al3+, on the Surface Adsorption and Self-Assembly of the Anionic Surfactant Alkyloxyethylene Sulfate and Anionic/Nonionic Surfactant Mixtures

The impact of multivalent counterions, Al(3+), on the surface adsorption and self-assembly of the anionic surfactant sodium dodecyl dioxyethylene sulfate, SLES, and the anionic/nonionic surfactant mixtures of SLES and monododecyl dodecaethylene glycol, C(12)E(12), has been investigated using neutron reflectivity, NR, and small angle neutron scattering, SANS. The addition of relatively low concentrations of Al(3+) counterions induces a transition from a monolayer to well-defined surface bilayer, trilayer, and multilayer structures in the adsorption of SLES at the air-water interface. The addition of the nonionic cosurfactant, C(12)E(12), partially inhibits the evolution in the surface structure from monolayer to multilayer interfacial structures. This surface phase behavior is strongly dependent upon the surfactant concentration, solution composition, and concentration of Al(3+) counterions. In solution, the addition of relatively low concentrations of Al(3+) ions promotes significant micellar growth in SLES and SLES/C(12)E(12) mixtures. At the higher counterion concentrations, there is a transition to lamellar structures and ultimately precipitation. The presence of the C(12)E(12) nonionic cosurfactant partially suppresses the aggregate growth. The surface and solution behaviors can be explained in terms of the strong binding of the Al(3+) ions to the SLES headgroup to form surfactant-ion complexes (trimers). These results provide direct evidence of the role of the nonionic cosurfactant in manipulating both the surface and solution behavior. The larger EO(12) headgroup of the C(12)E(12) provides a steric hindrance which disrupts and ultimately prevents the formation of the surfactant-ion complexes. The results provide an important insight into how multivalent counterions can be used to manipulate both solution self-assembly and surface properties.

Nanoblossoms: Light-Induced Conformational Changes of Cationic Polyelectrolyte Stars in the Presence of Multivalent Counterions

We analyze the structure of star-shaped polyelectrolytes in the presence of di- and trivalent counterions, and we use the gained knowledge to manipulate the polyelectrolyte's conformation by light. By applying dynamic light scattering and atomic force microscopy, we demonstrate that, at constant ionic strength, the arms of the cationic polyelectrolyte retract when adding multivalent counterions. Adding trivalent hexacyanocobaltate(III) ions leads to a collapse of the polyelectrolyte star even at low concentrations. This is shown by analysis of the star polyelectrolytes in solution as well as in the adsorbed state on mica surfaces. Considerably higher salt concentrations are necessary to obtain the same contraction of the polyelectrolyte star if the divalent tetracyanonickelate(II) ions are used. Sufficiently high multivalent counterion concentration leads finally to the precipitation of the polymer from the solution. We demonstrate that we can switch a polyelectrolyte star from the collapsed to the expanded state by transforming the trivalent hexacyanocobaltate(III) ions into a mixture of mono- and divalent ions by UV light. Thus, these collapsed stars react to light like "nanoblossoms". Moreover, polyelectrolyte stars precipitated through addition of the trivalent hexacyanocobaltate(III) ions can be redissolved by irradiation with light (photoinduced dissolution). Hence, the conformation and interaction of star polyelectrolytes can be switched by light. Possible applications of this novel way of manipulating polymers are discussed.

The EcoRI−DNA Complex as a Model for Investigating Protein−DNA Interactions by Atomic Force Microscopy

Atomic force microscopy (AFM) is a technique widely used to image protein-DNA complexes, and its application has now been extended to the measurements of protein-DNA binding constants and specificities. However, the spreading of the protein-DNA complexes on a flat substrate, generally mica, is required prior to AFM imaging. The influence of the surface on protein-DNA interactions is therefore an issue which needs to be addressed. For that purpose, the extensively studied EcoRI-DNA complex was investigated with the aim of providing quantitative information about the surface influence. The equilibrium binding constant of the complex was determined by AFM at both low and high ionic strengths and compared to electrophoretic mobility shift assay measurements (EMSA). In addition, the effect of the DNA length on dissociation of the protein from its specific site was analyzed. It turned out that the AFM measurements are similar to those obtained by EMSA at high ionic strengths. We then advance the idea that this effect is due to the high counterion concentration near the highly negatively charged mica surface. In addition, a dissociation of the complexes once they are adsorbed onto the surface was observed, which is weakly dependent on the ionic strength contrary to what occurs in solution. Finally, a two-step mechanism, which describes the adsorption of the EcoRI-DNA complexes on the surface, is proposed. This model could also be extended to other protein-DNA complexes.

DNA condensation and redissolution: interaction between overcharged DNA molecules

The effective DNA–DNA interaction force is calculated by computer simulations withexplicit tetravalent counterions and monovalent salt. For overcharged DNA molecules, theinteraction force shows a double-minimum structure. The positions and depths of theseminima are regulated by the counterion density in the bulk. Using two-dimensional latticesums and free energy perturbation theories, the coexisting phases for DNA bundles arecalculated. A DNA condensation and redissolution transition and a stable mesocrystal withan intermediate lattice constant for high counterion concentration are obtained.

Transport in doped conjugated polymers with polarons and bipolarons forming complexes with counter ions

In contrast to experimental evidence of bipolarons (BPs) in doped polymers, they have been ignored usually in both analyzing the electrochemical impedance spectra (EIS) and discussing the properties of organic devices. For EIS, the small-signal alternating current (ac) transport properties of polarons (P) and counter ions (C) are often modeled by transmission line equivalent circuits. Recently, this description has been extended to include steady-state effects and traps and, further, to account for the direct P–BP reaction. Here, the influence of traps and of the direct P–BP reaction are compared to each other. However, according to Kirova and Brazovskii [Synth. Metals 76 (1996) 229], the latter seems to be improbable, owing to the Coulomb repulsion except for the presence of a high counter ion concentration. They proposed a scenario with the formation of different complexes of P's and BPs with counter ions, and they suggested that the kinetics of the formation processes will be responsible, e.g., for hysteresis effects observed in organic devices. Here, the description is complemented by the first step, the formation of complexes from a polaron and a counter ion. The resulting small-signal ac equations are presented and conditions are analyzed, under which such complex formation processes can be important in EIS and in organic devices.

On the mobility of stiff polyelectrolytes

We present analytic calculations of the electrophoretic mobility of stiff charged molecules in solution, and results obtained by using different approximations for the hydrodynamic interactions are discussed. For low fields, where the average molecular conformation is hardly distorted from its Gaussian shape, we find an increase of the mobility with molecular size which, for chains up to 1000 segment lengths, follows approximately a power law with an exponent B. For sufficiently high counterion concentration (λD≲segment length) B is independent of the Debye length λD. Under the simplified assumption of spherical distributed counterions we find B to be proportional to the Debye length for λD larger than the segment length, except for the limits of Gaussian and rigid rod-type molecules. A brief comparison with numerical simulations is included.

Ion-pair LC of carbohydrates at alkaline pH. Separation and retention behaviour of glycoconjugates

A new technique for the separation of carbohydrates as ion-pairs in strongly alkaline solution is presented. Carbohydrates are weakly acidic and partly present as anions at pH≥12 [1]. They are retained as ion-pairs on polymeric solid phases (PRP-1 and PLRP-S) with a hydrophobic quaternary ammonium counter ion present in the mobile phase. The effects of nature and concentration of mobile phase components on the retention of carbohydrates have been investigated and an ion-pair distribution model is proposed. The influence of temperature indicated no changes in retention mechanism with high counter ion concentration, but the resolution decreased with increasing temperature. Saccharides added to the mobile phase were shown to increase the retention and the selectivity.

Lytic activity of monomeric and oligomeric melittin

The haemolytic activities of melittin and melittin tetramer as induced by high phosphate counterion concentration, were monitored. Monomeric melittin was found to be fully lytic, whilst tetrameric melittin lacked such activity. Under conditions where melittin was fully tetrameric attempts were made to covalently cross-link the native tetramer using a series of different chain length bifunctional imido esters. The cross-linked oligomers were fully lytic under conditions where melittin was demonstrated to lack such activity. This finding, together with molecular weight determinations and circular dichroism studies, indicated that the cross-linked melittin was quite different to the native tetramer. The haemolytic activity of melittin-containing solutions was related to the concentration of monomeric melittin. The effect of reduced dielectric constant (ϵ) on the aggregation behaviour of melittin and its derivatives was found to favour monomeric melittin.

Relaxation experiments in aqueous solutions of ionic micelles. 2. Experiments on the system water-sodium dodecyl sulfate-sodium perchlorate and their theoretical interpretation

Ionic micelles were studied as charged colloidal particles in order to interpret the qualitative difference between the theory of micellization kinetics and experimental results at high concentrations of surfactants. Results show that at low counterion concentration, the electrostatic repulsion prevents aggregation. At high counterion concentration, this reaction path is bypassed by a reversible aggregation, leading to increased rate of formation of proper micelles. The analysis yields an explanation for concentration dependence of the time constant for formation of nonionic micelles. 17 references.

Ion-pair chromatography of organic compounds

Ionized organic compounds can be extracted into organic phases as ion-pairs with counter ions of suitable hydrophobic character. The distribution ratio depends on the structure of the ion-pair and the concentration of the counter ion. Systems for liquid—liquid partition chromatography are easy to adapt to different types of compounds and have a high selectivity. The counter ion is selected on the basis of batch extraction procedures or chromatographic tests. A photometric detector can be used for samples of <0.1 nmole of non-absorbing compounds if a counter ion with high absorbance is used. Straight-phase systems with a high counter-ion concentration on a cellulose support have a high sample capacity and good separating efficiency ( H ⩽ 0.5 mm at a mobile phase speed of ⩽ 3 mm/sec when k′ ⩾ 5). Examples are given of straight-phase separations of carboxylic acids with a quaternary ammonium ion as counter ion in the stationary phase and amines and quaternary ammonium ions with picrate or β-naphthalene sulphonate as counter ions. Aminophenols have been separated by reversed-phase systems containing bis-(2-ethylhexyl)phosphoric acid in chloroform as the stationary phase.

Structural analysis of ion semi-permeable membranes by co-ion uptake and diffusion studies

Three different strongly basic exchangers with similar co-ion uptake behavior have very different co-ion diffusion characteristics. Not only are there differences in the order of magnitude of the co-ion permeabilities P at the same external electrolyte concentration c, but also marked differences in the shape of the P-c curves, which can be interpreted in terms of the internal micro- structure of the (inhomogeneous)membrand. Other structural details can be elucidated from the fact that polyvalent co-ions are more strongly excluded from local regions of high counterion concentration than are monovalent co-ions, and so lead to different P-c relations. Furthermore there was observed, especially for polyvalent ions, a great difference between the mechanisms of steady-state through-diffusion and of transient diffusion out of a saturated membrane, which also permits interpretation in terms of internal structure. The details obtained by the mathematical analysis of the diffusion data agree with, and go considerably beyond, what can be ascertained by electron microscopy. (auth)