Cross-linked polyvinylpyrrolidones with increased affinity and specificity for methyl orange and its homologs

Polyvinylpyrrolidones of various degrees of cross-linkage have been prepared by radical polymerization of N-vinylpyrrolidone with methylenebisacrylamide to regulate the fraction of cross-linkage. The insoluble polymers obtained were examined for their ability to bind methyl orange and its homologs, methyl orange, ethyl orange, propyl orange, and butyl orange at 5, 15, 25, and 35°C, respectively, in an aqueous solution. The first binding constants and the thermodynamic parameters that accompanied the binding were calculated. For any particular dye the extent of binding, the absolute magnitude of ΔF°, and the value of ΔS° increased as the degree of cross-linkage increased, starting with water-soluble polyvinylpyrrolidone (zero cross-linkage) and proceeding to the polymer with high cross-linking density. This behavior can be accounted for in terms of more extensive hydrophobic domains in the cross-linked polymeric matrix that enhances hydrophobic interactions in the binding process. Moreover, the cross-linked macromolecule polymerized in the presence of methyl orange and then stripped of the bound methyl orange shows substantially stronger binding for this small molecule than the polymer cross-linked in the absence of methyl orange. In contrast, the cross-linked polymer prepared similarly in the presence of the larger molecule, butyl orange, exhibits decreased affinity toward the smaller consolute, methyl orange, than either of the other polymers described. It seems, therefore, that the polymeric matrix provides favorable binding sites or pockets that can accommodate a specific small molecule. The preparative procedure, which uses a small-molecule template, molds into the polymer some structural specificity in the binding of small molecules.     

The binding of organic anions by polyvinylpyrrolidone: Determination of intrinsic binding constant and number of binding sites by competitive binding

The binding of 4′-dibutylaminoazobenzene-4-sulfonate (butyl orange, BO) and 1-amino-4-methylaminoanthraquinone-2-sulfonate (AQ) by crosslinked polyvinylpyrrolidone and the competition between BO and AQ for binding sites of the polymer were examined. The equilibrium data showed competitive binding. Thus the intrinsic binding constants and the number of binding sites can be evaluated easily and precisely by competitive binding. The thermodynamic parameters that accompained the binding were calculated from the intrinsic binding constant and its temperature-dependence. The thermodynamic data for BO showed that the binding process is athermal and stabilized entirely by the entropy term. The binding of BO to the polymer is entropically favorable as a result of the operation of the hydrophobic effect. In contrast, with AQ a favorable free energy for a dye-polymer complex formation is associated with a large negative enthalpy and a small entropy. Therefore it is likely that the binding of AQ occurs by energetic forces and that the large aromatic ring of the dye contributes to the binding energy. On the average, BO and AQ can bind to the crosslinked polymer to the extent of 1 dye/ca 73 basemol in 0.1M tris-acetate buffer, pH 7.0.     

Interaction of polyvinylpyrrolidone with methyl orange and its homologs in aqueous solution: Thermodynamics of the binding equilibria and their temperature dependences

The interaction of polyvinylpyrrolidone with methyl orange, ethyl orange, propyl orange, and butyl orange has been studied by an equilibrium dialysis method at 5, 15, 25, and 35°C. The first binding constants and the thermodynamic parameters in the course of the binding have been calculated. It was found that the free energy and the enthalpy changes are all negative and the entropy change is largely positive. The longer the alkyl chain of the dyes, the more positive is the enthalpy change (though it is always in the negative direction) and hence the larger is the entropy change. The favorable free energy of the binding of butyl orange observed for the formation of the dye–polymer complex seems to be a result of a favorable entropy change rather than any favorable enthalpy change. Temperature dependences of the thermodynamic functions were apparently observed. That is, ΔF and ΔH become larger in absolute magnitude as the temperature increases. The positive quantity of ΔS tends to decrease with increasing temperture. All these facts obtained can be interpreted satisfactorily by the hydrophobic interaction between hydrocarbon portions of the dyes and nonpolar parts of the macromolecule.     

Interaction of poly(vinylpyrrolidone) with methyl orange and its homologs in aqueous solution over the temperature range 60–90°c

The binding of methyl orange, ethyl orange, propyl orange, and butyl orange by poly(vinylpyrrolidone) has been examined by a technique of equilibrium dialysis over a high temperature range (60–90°C). The first binding constants and the thermodynamic parameters in the course of the binding were evaluated. The results obtained at these temperatures were compared to those at lower ones (5–35°C) described previously in order to estimate the contribution of hydrophobic bonds to the binding. It was found that at the 60–90°C range complex formation between the dye and the macromolecule is associated with an exothermic enthalpy change and a positive entropy change. The enthalpy and entropy changes of the binding are of the order of ?4.5 kcal/mole and 6 eu, respectively, for each dye measured. Thus the binding is mainly enthalpy-controlled. Furthermore the effect of the alkyl chain length of the dye on both the ΔH° and ΔS° values is not pronounced. Also temperature dependences of the ΔH° and ΔS° terms were not observed. All these observations in the higher temperature range can be explained as a result of the disruption of water structure in the binding environment and hence a decrease in hydrophobic bond formation between the dye and the polymer.     

Thermodynamics of binding of methyl orange and its homologs by poly(vinylpyrrolidone): The effect of inorganic electrolytes

The extent of binding of methyl orange, ethyl orange, propyl orange, and butyl orange by poly(vinylpyrrolidone) has been measured in aqueous solutions of inorganic electrolytes such as NaCl, LiCl, NaSCN, and NaClO₄ by an equilibrium dialysis method. The effect of the salts on the first binding constants and the thermodynamic functions which are accompanied by the dye—polymer association process was investigated relative to the corresponding values in the absence of such salts. It was found that in aqueous solutions of NaCl and LiCl the enthalpy change accompanying the binding is small and the largest contribution to the free energy of binding is from the positive entropy gain. For NaSCN and NaClO₄, the values of ΔF° and ΔH° were both large and negative and the value of ΔS° was small and negative. Thus, the favorable free energy for the complex formation was due entirely to the negative enthalpy term. These characteristics of the thermodynamic quantities are discussed in terms of changes in structural properties of water in the vicinity of the binding entities and conformational changes of the polymer to which the dye is bound due to the added foreign electrolytes.     

Peculiar temperature dependence of the binding of methyl orange and its homologs by 2-diethylaminoethyl methacrylate-N-vinyl-2-pyrrolidone copolymers

2-Diethylaminoethyl methacrylate (DEAEMA)–N-vinyl-2-pyrrolidone (VPy) copolymers of various compositions have been synthesized. The resultant copolymers were examined for their ability to bind methyl orange and its homologs, in particular butyl orange, at 5, 15, 25, and 35°C in aqueous solutions. The amount of binding of butyl orange is much higher with the copolymers than with polyvinylpyrrolidone or with 2-hydroxyethyl methacrylate–N-vinyl-2-pyrrolidone copolymers. Introduction of only 3% of the hydrophobic DEAEMA residue increases markedly the binding affinity toward the cosolute. Maximal binding is obtained at 15°C in the temperature range measured. This peculiar temperature dependence of the extent of binding is explicable on the basis of hydrophobic effects involved in this binding. The peculiar temperature dependence disappeared in aqueous solution of NaSCN which acts as a water-structure breaker: the extent of binding changes regularly with temperature. This is interpretable only in terms of reduction of hydrophobic contribution to the binding. With propyl orange, which is a less hydrophobic cosolute than butyl orange, the peculiarity of the binding was not detected.     

Binding of methyl orange and its homologs by powdered nylon 612: Peculiar temperature dependence on the binding

The ability of powdered Nylon 612 to bind methyl orange, ethyl orange, propyl orange, and butyl orange was investigated at 5, 15, 25 and 35°C in an aqueous solution. The amount of binding of the dye is much higher with this polyamide than with powdered Nylon 66 reported previously,¹ although the former polymer has fewer amide end groups. The Van't Hoff plots of the first binding constant for the binding of butyl orange and propyl orange by powdered Nylon 612 exhibit a bell-shaped curve, whereas the plots for methyl orange and ethyl orange do not. Maximal binding occurs at approximately 15°C for propyl orange and at about 25°C for butyl orange. This is the first instance where the peculiar temperature dependence of the binding constant has been found in the binding of propyl orange, whose hydrophobicity is less than that of butyl orange. These tendencies can be accounted for in terms of increased hydrophobic of butyl orange. These tendencies can be accounted for in terms of increased hydrophobic domains in powdered Nylon 612 and enhanced hydrophobic contributions in the binding process.     

Temperature-responsive swelling and deswelling of the copolymers from vinyl ether of ethylene glycol and butyl vinyl ether

Linear and crosslinked copolymers of a vinyl ether of ethylene glycol (2-hydroxyethyl vinyl ether, ( 1 )) and butyl vinyl ether ( 2 ) are synthesized by α-irradiation polymerization. It is shown that the linear copolymers exhibit a phase separation phenomenon in dependence of the temperature due to the destruction of hydrogen bonds and the enhancement of hydrophobic interaction in aqueous solution. The processes of reversible swelling or shrinking upon temperature change are demonstrated for polymer networks.     

Interaction of crosslinked polyethylenimine with a homologous series of methyl orange derivatives in aqueous solution

Polyethylenimine (PEI) was crosslinked with dichloroethane, glyoxal, or glutaraldehyde and polymers of various degrees of crosslinkage were made. The insoluble polymers obtained were examined for their ability to bind methyl orange and its homologs, methyl, ethyl, propyl, and butyl orange at 5, 15, 25, and 35°C, respectively, in an aqueous solution. PEI crosslinked with glutaraldehyde showed markedly increased binding affinity toward these cosolutes compared with the polymers crosslinked with dichloroethane or glyoxal. The extent of the binding increased with an increase in the degree of crosslinkage. These results suggest that the enhancement of the binding by the crosslinking is due mainly to a dual effect, introduction of hydrophobic moieties and proximity of neighboring polymer chains. The first binding constants and the thermodynamic parameters that accompanied the binding were calculated. The thermodynamic data show that the binding process is athermal and is stabilized entirely by the entropy term. Water-soluble PEI exhibited stronger cooperative interactions than the crosslinked polymer because the mobilities of the chains of the former are greater than those of the latter.     

Peculiar temperature dependence on the binding of methyl orange and its homologs by 2-hydroxyethyl methacrylate-N-vinyl-2-pyrrolidone copolymers

2-Hydroxyethyl methacrylate (HEMA)-N-vinyl-2-pyrrolidone (VPy) copolymers of various compositions have been prepared. The copolymers obtained were examined for their ability to bind a homologous series of methyl orange derivatives, methyl orange, ethyl orange, propyl orange, and butyl orange, at 5, 15, 25, and 35°C, respectively, in an aqueous solution. The first binding constants and the thermodynamic parameters that accompanied the binding were evaluated. The binding ability of the copolymer for the small cosolute was enhanced with an increase of the HEMA content in the copolymer. Moreover, a bell-shaped curve appeared in the binding of butyl orange by the copolymers having higher HEMA residues when the first binding constant was plotted as a function of temperature, whereas no such phenomenon was detected for the copolymers with less HEMA content or for the less hydrophobic dye, methyl orange, ethyl orange, or propyl orange. This peculiar temperature dependence of the first binding constant shows that the enthalpy of the binding varies from a positive (unfavorable) value below ca. 15°C to a negative (favorable) one above this temperature. This behavior can be accounted for in terms of more hydrophobic effects involved in the binding process.     

Binding of methyl orange and its homologs by powdered nylon 66

Powdered Nylon 66 was prepared as a model of amorphous polymers. The resultant powder polyamide was composed of only amorphous regions. The extent of uptake of the acid azo dyes, a homologous series of methyl orange derivatives, by the polymer was measured in an aqueous solution. The first binding constants and the thermodynamic parameters in the course of the binding were evaluated. The thermodynamic behaviors obtained are very similar to those of crosslinked polyvinylpyrrolidone. The favorable free energy of the binding is accompanied by an entropy gain and an exothermic enthalpy change. The shorter the alkyl chain of the dyes, the more negative is the enthalpy change and, hence, the smaller is the entropy change. The thermodynamic data for butyl orange showed that the binding process is athermal and is wholly an entropic effect. The binding of the dyes to the matrix is entropically favorable as a result of the operation of the hydrophobic effect. In addition, an electrostatic force is operative between the sulfonate group on the dyes and the terminal amino groups on the polyamide.     

Temperature dependence on the binding of butyl orange by bovine serum albumin

The binding of 4′-dibutylaminoazobenzene-4-sulfonate anion (butyl orange) by bovine serum albumin has been examined quantitatively by an equilibrium dialysis method at 5, 10, 15, 20, 25, and 35°C. The first binding constants and the thermodynamic parameters for the formation of the first dye anion-protein complex have been calculated. The peculiar temperature dependence of the first binding constant could be observed. That is, the value of the first binding constant increases with increasing temperature until it reaches a maximum value at approximately 18°C and then decreases with raising temperature. Accordingly, this binding process is exothermic above 18°C and is endothermic below 18°C. Near 18°C the process exhibits athermal reaction. From the thermodynamic data obtained, it is evident that the favorable free energy of the binding is accompanied by an entropy gain and that the enthalpies of the binding vary from a positive (unfavorable) value below 18°C to a negative (favorable) one above 18°C. Furthermore an apparent temperature dependence of the thermodynamic functions was observed. That is, ΔF° becomes larger in absolute magnitude as the temperature increases. The positive quantity of ΔS° tends to decrease with increasing temperature. All these facts can be interpreted satisfactorily in terms of hydrophobic interactions between hydrophobic portions of the dye and nonpolar parts of the albumin.     

Thermodynamics of binding methyl orange and its homologs by polyvinylpyrrolidone: The effect of hexamethylenetetramine

Hexamethylenetetramine (HMT) has been examined for its effect on the binding of methyl orange homologs, methyl orange, ethyl orange, propyl orange, and butyl orange by polyvinylpyrrolidone (PVP). In the presence of HMT the entropy changes associated with the binding tend to become more positive and the absolute magnitude in the enthalpy changes becomes smaller compared with those in the absence of HMT. These tendencies are accounted for in terms of the water-structure-promoting effect of HMT, hence the enhancement of hydrophobic interactions in the binding. PVP undergoes changes in conformation on the addition of HMT and its conformation becomes more compact. This also increases the contribution of the hydrophobic interactions to the macromolecule-small molecule interaction. Some other effects exerted by the added HMT on the binding system are also described.     

Spectral Studies on the Binding Behavior of Cationic Dyes and Surfactants with Bacterial Polysaccharide of Klebsiella K43

The binding behavior of acidic capsular polysaccharide (SPS), isolated from Klebsiella serotype K43, with oppositely charged dyes and surfactants have been studied by way of absorbance and emission spectroscopic measurements. Each repeating unit of the SPS consists of three D-mannose, one D-galactose, and one D-glucuronic acid residue. The anionic polysaccharide exhibited chromotropic character and induced strong metachromasy in the cationic dye, pinacyanol chloride (PCYN) through the formation of a 1:1 stoichiometric complex. Evaluation of thermodynamic parameters, viz., changes in free energy (ΔG), enthalpy (ΔH), and entropy (ΔS), for the formation of dye-polymer complex and studies on the effect of different cosolvents were also evaluated to shed light on the binding nature as well as the extent of stability of the dye-polymer complex. Fluorescence of the cationic dye acridine orange (AO) was quenched with the progressive addition of SPS, which was found to be of Stern-Volmer type. Cationic surfactants in their pure form as well as in the mixed state with nonionic surfactant (Tween-20), replaced the dye bound to the polymer matrices; thus the original band intensities of the dyes could be reverted. Such studies revealed the involvement of both electrostatic as well as hydrophobic interactions between the dye-polymer as well as surfactant-polymer aggregates.     

Equilibrium study of polymer–polymer complexation of poly(methacrylic acid) and poly(acrylic acid) with complementary polymers through cooperative hydrogen bonding

The degree of linkage, θ, defined as the ratio of the binding groups to the total of potentially interacting groups and the stability constant K of the polymer–polymer complexes in the systems poly(methacrylic acid)–poly(ethylene glycol), poly(acrylic acid)–poly(ethylene glycol), and poly-(methacrylic acid)–poly(vinyl pyrrolidone) in aqueous and aqueous alcohol media were determined as a function of temperature by potentiometric titration. It was found that θ and K are strongly dependent on chain length, temperature, and medium and that hydrophobic interaction is a significant factor in the stabilization of the complexes. The enthalpy and entropy changes and the cooperativeness parameter of the systems were calculated. A mechanism for the complexation in terms of cooperative interaction was proposed.     

Solvation of amides of carboxylic acids in aqueous solutions of ethylene glycol

Enthalpies of solution of amides of formic, acetic, and propionic acids with different degrees of N-substitution in aqueous solutions of ethylene glycol were measured at 298.15 K. The concentration of ethylene glycol did not exceed 4 mol kg^–1. The reasons for increasing endothermic values of the enthalpies characterizing the amide transfer from water to a mixed aqueous-organic solvent on going from primary to tertiary amides and from formamides to the corresponding acetamides are discussed. The enthalpic coefficients of pair interactions between amides and ethylene glycol in water were calculated. The endothermicity of the interaction of the alkyl groups of the amide molecules with ethylene glycol results in positive values of the coefficients. The coefficient values increase with the enhancement of the hydrophobic properties of hydrophilic non-electrolytes (urea, formamide, ethylene glycol) due to an increase in the contribution of the hydrophobic component and a decrease in the contribution from the interaction of the polar groups of amides to the total interaction.     

Affinity partitioning of enzymes using dextran-bound procion yellow HE-3G. Influence of dye-ligand density

The dye Procion Yellow HE-3G was bound to dextran of molecular weight 70,000 and the partitioning of this dye-polymer within an aqueous two-phase system containing dextran and poly(ethylene glycol) was studied as function of ligand density, polymer concentration, type of salt, concentration of salt and concentration of dye-dextran. Even moderate dye:dextran molar ratios (5-8) make the partitioning strongly salt-dependent. The dye-dextran can be directed to either the upper or the lower phase with partition coefficients from 0.02 to 28 by using salts. The dye-dextran in the two-phase system affects the partitioning of dye-binding enzymes (lactate dehydrogenase, glucose-6-phosphate dehydrogenase, 3-phosphoglycerate kinase) towards the dye-containing phase. Measurements of competition with nucleotide binding show an increased affinity of the dye for the enzyme with increasing ligand:dextran ratio. Theoretical considerations indicate that 1-2 dextran molecules are attached per enzyme molecule when affinity partitioning is fully developed.     

Binding methyl orange and hydrophobic fluorescent probes by poly(2-dimethylaminoethyl methacrylate) and copolymers of 2-dimethylaminoethyl methacrylate and N-vinyl-2-pyrrolidone: pH dependence of the binding and fluorescence intensity

The pH dependence of the interaction of poly(2-dimethylaminoethyl methacrylate) and copolymers of 2-dimethylaminoethyl methacrylate and N-vinyl-2-pyrrolidone with methyl orange, 2-p-toluidinylnaphthalene-6-sulfonate (TNS), and 1,6-diphenyl-1,3,5-hexatriene (DHT) was studied by equilibrium dialysis and fluorescence measurements at pH's 7–10. The first binding constant accompanying the binding of methyl orange and TNS by the polymers, in particular the homopolymer, shows a maximum around pH 8 and maximal fluorescence intensity of TNS is obtained around pH 8.5 in the presence of the polymers. To elucidate these observations the pH-induced conformational changes of the homopolymer were examined by potentiometric titration and viscosity measurements and the thermodynamic parameters that accompany the binding were calculated. The polymer was found to change from an extended coil at lower pH to a compact coil at higher pH. The electrostatic attraction between the sulfonate group of the small molecule and the protonated nitrogen atoms on the polymer is increased at lower pH and the hydrophobic interaction between the hydrophobic moieties of the polymer and the small molecule is enhanced at higher pH. The results obtained for the dye binding and fluorescence intensity were discussed in terms of the electrostatic and hydrophobic interactions.     

Effect of copolymer composition on interpolymer complex formation of (co)poly(vinyl ether)s with poly(acrylic acid) in aqueous and organic solutions

The complex formation between vinyl ether of poly(ethylene glycol‐co‐vinyl butyl ether) with poly(acrylic acid) has been considered in aqueous and isopropanol solutions. The effect of copolymer composition on the complex formation process was clarified. It has been shown that the incorporation of hydrophobic fragments into macromolecules enhances the hydrophobic stabilization of polycomplexes in aqueous solutions. In organic media this effect disappears. The stability of polycomplexes formed both in aqueous and in organic solutions in respect to the addition of dimethylformamide has been studied.     

Amphiphilic copolymers with increased affinity for substrates

The copolymers of methyl quaternized 2-dimethylaminoethyl acrylate and styrene, 2-vinyl naphthalene, acrylic acid iso-octyl ester, or acrylic acid n-butyl ester have been prepared. Studies were made of the binding of a “binding probe,” methyl orange, by the copolymers in aqueous solution. The first binding constants and the thermodynamic parameters in the course of the binding were evaluated. Furthermore, the intensity of fluorescence of a hydrophobic fluorescent probe, 2-p-toluidinylnaphthalene-6-sulfonate, in the presence of these polymers was investigated. In addition, the fluorescence spectra of monomer and excimer emissions of the polymers with aromatic residues were measured. The excimer/monomer fluorescence intensity ratio was studied in the presence of various additives such as methyl orange, urea, methanol, and NaCl to gain an insight into the nature of microdomains in the polymer. The nature and phenomena of dye binding and hydrophobic fluorescent probe binding with the polymers are discussed. © 1993 John Wiley & Sons, Inc.