John B. Lawton

Polyelectrolyte complexes. 2. Interaction between collagen and polyanions

Abstract The electrostatic interactions that occur in connective tissue between polyanions and proteins have been studied in model systems by a technique involving a fluorescent probe, acridine orange. It was found that collagen bound more strongly than bovine serum albumin to the polyanions studied. At pH 3.0, collagen formed strong complexes of definite stoichiometry with chondroitin-4-sulphate, chondroitin-6-sulphate, heparin and polystyrene sulphonate that were stable in sodium chloride solution of 0.1 M. The complexes of collagen with hyaluronic acid, or carboxymethylcellulose were less stable. The effect of pH variations (3.0–9.0) on the binding was investigated. Critical electrolyte concentrations (NaCl) were determined for complexes of collagen with glycosaminoglycans that dissociated at salt concentrations below that at which collagen precipitates. The values obtained were, 0.1 M for hyaluronic acid, and ∼0.5 M for chondroitin sulphate.

Calorimetric determination of the enthalpy change for the binding of methyl orange to poly(vinylpyrrolidone) in aqueous solution

Abstract Thermodynamic parameters have been determined for the interaction of methyl orange and poly(vinylpyrrolidone) in aqueous solutions containing tris, tris and hydrochloric acid, or water only. Enthalpy changes were determined by use of a flow microcalorimeter. Free energy changes were calculated from the results of equilbrium dialysis studies. The results are interpreted in terms of the behaviour of poly(vinylpyrrolidone) in these systems. Analysis of data shows that the hydrophobic interactions in the systems are in the order: tris > wateronly > tris-HC1.

Fluorescence polarization studies of the interactions between protamine and enzymes in aqueous solutions

Abstract Fluorescence polarization has been used to study the interaction of dansylated protamine with the enzymes: pepsin, α-chymotrypsin, alkaline phosphatase and invertase. These interactions have been compared with those between dansylated protamine and polyacrylate, or polyvinylsulphate. Each of the various complexes was found to be dissociated by the addition of sodium nitrate and a critical electrolyte concentration (CEC) was determined for each system, to allow assessment of the relative order of binding to the dansylated protamine. This order was: polyvinylsulphate >pepsin >polyacrylate >alkaline phosphatase >α-chymotrypsin. The strength of binding was also assessed by determination of a binding constant, Ka. The values of Ka showed the same relative order of binding as the CEC values. Invertase behaved similarly to the other enzymes, but it was not possible to obtain an unambiguous assessment of the comparative strength of binding. In each case, the stoichiometry of the complex was also determined.

Interaction of acridine orange and polyanions: fluorimetric determination of binding strengths and the influence of simple electrolytes

Binding affinities of Acridine Orange and six polyanions have been determined by measuring association constants in essentially salt-free solution, and also by observing competition between the dye and a simple salt for the polyanion site. Differences were found in the dye-binding order for the six polyanions using the two techniques. These differences were related to the breakdown in the uniform binding behaviour of carboxy and sulphate groups bound to the same polymer backbone as the salt concentration is increased. A mechanism is proposed which fits the thermodynamic parameters calculated by the application of Schwarz treatment and the Langmuir isotherm. The difference in the free-energy of binding between the strongest binding polymer, polystyrene sulphonate (ΔG°=–41 kJ mol–1) and the weakest binding, hyaluronic acid (ΔG°=–29 kJ mol–1) could be accounted for by considering coulombic site-dye interactions which destabilise the dye aggregates. The main influence promoting dye binding appears to be the free-energy of dye aggregation.The degree of co-operativity of dye binding, q, has also been determined. An attempt has been made to relate this parameter to the physical properties of the polyanion chain. It appears unlikely that q has the significance initially attached to it.

Methylation as a means of location of anionic groups in some cellulose phosphates

Abstract The distribution of methyl groups after methylation of celluloses containing phosphate groups is quite different from that obtained after methylation of cellulose. The results indicate a probable high level of phosphorylation at positions 2 and 6 of d -glucopyranosyl units, and give supporting evidence for the recognised fibrillar models for cotton.

Modification of the surface characteristics of cotton by the introduction of anionic groups, and the mechanism of ion binding

The introduction of carboxymethyl, phosphate and sulfate groups into cotton cellulose considerably modifies the capacity of the cellulose to bind cationic systems. The ion-binding characteristics of these anionic derivatives have been quantitatively studied using the cationic dye methylene blue. Diffuse reflectance spectral measurements demonstrate that two types of sulfated and phosphorylated celluloses can be produced, which differ considerably in the interaction of their anionic groups with the dye. Anionic systems are, in contrast, repelled by the anionic substituents. In this way the resistance of the cotton cellulose to wet soiling can be considerably increased. The mechanism of ion binding to anionic celluloses has been examined and the information utilized to predict methods of modifying the surface characteristics of cotton cellulosee.

Calorimetric determination of the enthalpy change for the binding of xanthene dyes to polycations in aqueous solutions

Abstract Thermodynamic parameters have been determined for the interaction of two xanthene dyes, fluorescein (Fl) and eosin Y (Eo-Y) with the polycation (poly(paraxylylviologen dichloride)) (PPXVCl 2 ) and for the interaction of Fl with poly(4-vinyl pyridinium methyl chloride) (P4VPMeCl) in aqueous solutions. Enthalpy changes were determined by the use of a flow microcalorimeter. Free energy changes were calculated from the results of spectrophotometric studies. The results have been interpreted in terms of the structures of the dyes and the conformations of the polycations. Analysis of the data shows that the electrostatic interactions in these dye-polycation systems are in the order: P4VPMeCl > PPXVCl 2 for the polycations, and Eo-Y > Fl, for the two dyes. The free energy changes for the dye-polycation interactions are in the order: Fl-P4VPMeCl > Eo-Y-PPXVCl 2 > Fl-PPXVCl 2 .

Polyanions and their complexes. Part 10. Mechanism of dye binding to chemically different sites

Thermodynamic parameters have been determined for the ion-exchange of Methylene Blue cations with sodium ions or potassium ions on cellulosic polyanions. These polyanions resemble biological oolyelectrolytes with respect to their ion binding characteristics. The chemical nature of the site determines the free energy change for the binding of Methylene Blue in the order SO3– > CO2– > PO2– > CH2CO2–. The mechanism of dye binding is discussed and an explanation is advanced for the influence of the site on the order of binding. It is shown that the affinity of cationic dyes for a polyanion may be predicted from a knowledge of their tendency to aggregate in aqueous solution.