The suggestion is made that, in solution, the flexible-chain molecules of dextran can undergo an osmotic compression as concentration is increased. Approaches are developed described the molecular shrinkage (i) as arising from intra- and inter-molecular forces, (ii) based on the molecular characteristics of the dextran, and (iii) as estimated by viscosity measurements. Comparison with the macroscopic shrinkage of cross-linked dextran (Sephadex) beads [Edmond, Farquhar, Dunstone & Ogston (1968) Biochem. J. 108, 755-763] is made. In all systems studied, the experimental estimates of compression, both from gel-shrinkage and viscosity measurements were in reasonable agreement with theoretical predictions. The interpretation of the viscosity concentration-dependence was applied to compact structures (albumin and Percoll). Their behaviour was in marked contrast with that of dextran. It is noted that molecular compression may be important in considering transport processes in and thermodynamic properties of concentrated systems.

A theoretical expression is derived for concentration dependence of elution volume in the gel chromatography of a non-interacting solute. Experimental results for bovine serum albumin on Sephadex G-100 are shown to be in good agreement with the predicted gel-chromatographic behaviour. The theoretical treatment of concentration dependence is extended to include a solute undergoing rapid reversible polymerization (nA in equilibrium C). Computer simulation of gel-chromatographic data for monomer-dimer systems on Sephadex G-100 is used to illustrate the deficiencies of earlier empirical approaches, and also the potential of the present treatment, of allowing for non-chemical concentration dependence in gel chromatography of polymerizing solutes.

1. A bilayer strip, cut from a thin layer of cross-linked polyacrylamide gel cast on to cellulose tissue, forms an open circular loop whose ends are close together. Shrinkage of the gel, in response to the osmotic pressure of a non-penetrating solution, causes a proportional separation of the ends of the loop. This is measured with a microscope and micrometer eyepiece. 2. The resulting effective sensitivity is about 30 times that of the Sephadex-bead osmometer (Ogston & Wells, 1970), i.e. of the order of 5Pa, comparable with that of a membrane osmometer. Use of gel up to 70% (w/v) allows the measurement of molecular weights, as low as 1500 in favourable cases, with an accuracy of 1–2%. The useful range of osmotic pressure is up to 5kPa. A single measurement requires 0.5ml of solution. Equilibration is completed in 20–30min. 3. The method is illustrated by measurements on human serum albumin, ovalbumin, cytochrome c , samples of dextrans, polyvinyl alcohol, and polyethylene glycols 6000 and 1000.

1. Observations are reported on the variation of the swelling of sulphoethyl-Sephadex C-50 and C-25, and of the partition of NaCl between solution and gel, with the concentration of NaCl. 2. The results were interpreted in terms of Manning's (1969) treatment of the interactions between polyionic polymers and simple electrolytes and of Flory's (1953) treatment of the swelling of gels. 3. The results indicate net inner osmotic pressures as high as 3.6×10 5 Pa (140 mosmolar=3.6×10 5 N/m 2 ) in 0.15 m -NaCl. 4. It is suggested that cartilage may have net inner osmotic pressure of the order of 10 5 Pa.

1. The measurement of osmotic pressure by means of a single bead of Sephadex (Edmond, Farquhar, Dunstone & Ogston, 1968) has been made more precise by immobilizing the bead on a fine needle. The design, calibration and use of the osmometer are described. 2. The method is particularly suitable for measuring high osmotic pressures in solutions of high-molecular-weight solutes, which must not penetrate the Sephadex to a significant extent. 3. With Sephadex G-50 the limit of precision is about 1.5cmH 2 O and the lower limit of molecular weight for a solute of compact molecular form is about 10 5 . 4. The time required for each equilibration is less than 10min. 5. An impaled bead can be stored in the dry state without affecting its calibration. 6. Measurements on a sample of polyvinyl alcohol, degree of polymerization stated as 1750±50, gave ¯m n 43000±3000 and A 2 (6.0±0.24)×10 −4 .

(1) We have measured the incompatible phase separation that occurs in a polyethylene glycol–sodium dextran sulphate–sodium chloride–water system and have determined a critical point. (2) We have measured the activity coefficients of sodium chloride in critical-point concentrations of polyethylene glycol and sodium dextran sulphate respectively, and the osmotic coefficient of sodium dextran sulphate at the critical-point concentration. (3) With use of the relevant thermodynamic equations for a quaternary ionic system, we have determined the interaction coefficients between polyethylene glycol and dextran sulphate and between polyethylene glycol and sodium chloride. The former could be due mainly to volume exclusion, but the latter is too large to be explained on that basis.

1. We have measured the partition coefficients of bovine serum albumin with Sephadex grades G-100, G-150 and G-200, and of a dextran ([unk] n 19700) and a polyethylene glycol ([unk] n 8000) with Sephadex G-200. We have also measured the effects of these solutes on the inner volumes of the grades of Sephadex. 2. The results can be described with fair consistency by means of a simple thermodynamic treatment that makes use of the virial coefficients of Sephadex and of the solute, and of a coefficient that expresses their interaction. This coefficient is related to the ‘exclusion volume’ of Sephadex for the solutes. 3. The Sephadex G-200–polyethylene glycol system shows anomalies of behaviour that are ascribed to the occurrence of ‘incompatible’ phase separation within the Sephadex beads.

1. Simple thermodynamic expressions are used to describe the properties of uncharged binary and ternary polymer solutions, in particular the sedimentation equilibrium of binary systems and the osmotic pressures and ‘incompatible’ phase separations of ternary systems. 2. Sedimentation-equilibrium experiments were performed on four samples of dextran and two of polyethylene glycol. The critical points of the phase diagrams were determined for the mixed solutions of polyethylene glycol–dextran–water and of polyethylene glycol–bovine serum albumin–0·2 m -sodium chloride solution. Osmotic pressures were measured on a single-phase mixed solution of a polyethylene glycol and a dextran. By use of the simple thermodynamic expressions consistent values of second virial and interaction coefficients for the materials used were obtained from these experiments. 3. The interpretation of the values of the second virial and interaction coefficients, on the basis of three models of molecular interaction, is discussed.

1. Hyaluronic acid was isolated from ox synovial fluid by sedimentation equilibrium in a caesium chloride density gradient (Silpananta, Dunstone & Ogston, 1967). The product was almost free from chondroitin sulphate and from protein. 2. Its composition did not differ significantly from that of the carbohydrate part of the protein-containing material isolated by filtration. Its physicochemical properties and molecular configuration were similar, except for its viscosity, which showed markedly reduced concentration-dependence and shear-dependence. This suggests that the associated protein tends to form links between molecules of hyaluronic acid. 3. The accurate measurement of viscosity at very low velocity gradient, by use of the damping of oscillations in a Couette viscometer, is described. 4. A method is described for measuring, approximately, the thermodynamic non-ideality of a solute from the shape of its schlieren curve at sedimentation equilibrium in a density gradient. 5. A value for the partial specific volume of hyaluronic acid in dilute salt solution was calculated from its isopycnic density in a caesium chloride gradient.

1. Sephadex in bead form shows reversible changes of inner volume when immersed in solutions of a non-penetrating solute. These changes are in accordance with the theory of Flory (1953) for the swelling of gels. This makes possible the use of single beads for measuring the osmotic pressures of polymer solutions, up to or beyond 1kg./cm. 2 . 2. Measurements of the inner volume of Sephadex G-200 by equilibrium dilution, at various concentrations of dextran 500, gave values in agreement with those obtained from the dimensions of single beads. 3. Measurements by chromatography of the inner volume of Sephadex G-200 gave values that differed slightly but significantly from those obtained by the other methods. Similar small disagreements were found between the dilution and chromatographic values for bovine serum albumin measured in the presence of various concentrations of dextran 500. The major factor that affects the dependence of the chromatographic elution volume (of dextran 500) on concentration is shown to be the change of the inner volume of the Sephadex, with dynamic factors playing a minor role. 4. Contrary to the findings of Ackers (1964), the equilibrium dilution and chromatographic methods gave closely agreeing values for the distribution coefficient, K av. , of bovine serum albumin on Sephadex G-200; this cast doubt on his explanation of the nature of chromatographic separations on Sephadex.