Diffusion of oxygen in water and hydrocarbons using an electron spin resonance spin-label technique.

The Smoluchowski equation for the bimolecular collision rate of dissolved oxygen molecules with spin labels yielded values for the diffusion constant of oxygen in water that are in agreement with the Stokes-Einstein equation (D infinity T/eta, where eta is the macroscopic viscosity) and with published values obtained by conventional methods. Heisenberg exchange at an interaction distance of 4.5 A occurs with a probability close to one for each encounter. In mixed hydrocarbons (olive oil, paraffin oils) and sec-butyl benzene, D infinity (T/eta)rho, where rho lies between 0.5 and 1. Oxygen diffuses in the hydrocarbons between 10 and 100 times more rapidly than predicted from the macroscopic viscosity. Similar results would be expected for diffusion of oxygen in model and biological membranes. Parallel measurements of rotational diffusion of the spin labels show little correlation with measurements of translational diffusion of oxygen. Dipolar interactions between spin labels and oxygen appear negligible except in the limit of highest viscosities.     

Kinetics of partly diffusion controlled reactions. VII — Pyrene excimer formation in erythrocyte membranes

Intermolecular excimer forming systems such as pyrene have been proposed as probes for the microviscosity measurement of model and biological membranes. However, our experimental study of pyrene excimer formation in erythrocyte membranes and the theoretical calculations show that it is quite difficult to get the diffusion coefficient of pyrene in membranes and therefore the microviscosity of its environment.     

c,T-dependence of the viscosity and the self-diffusion coefficients in some aqueous carbohydrate solutions

Self-diffusion coefficients for both components are reported for the highly concentrated aqueous solutions of some disaccharides and fructose as a function of temperature and concentration. These data are complemented by viscosity measurements. The disaccharides studied are sucrose, alpha,alpha-trehalose, allosucrose, and leucrose. Up to a sugar concentration of approximately 30% w/w the viscosity and the self diffusion coefficients of the four disaccharides are identical within experimental error for a given concentration and temperature. Water diffusion shows no differences in the four systems studied under these conditions. At higher concentrations significant differences are observed that become more pronounced with increasing temperature. Analysis of the data by the VTF equation yields the result that at a given concentration the self diffusion coefficients of the sugar Dc and the viscosity eta are described by identical ideal glass transition temperatures T0, while the diffusion of the water D(W) molecule decouples from these properties. T0(W) is always lower than T0(c,eta).     

Probe diffusion in aqueous poly(vinyl alcohol) solutions studied by fluorescence correlation spectroscopy

We report fluorescence correlation spectroscopy measurements of the translational diffusion coefficient of various probe particles in dilute and semidilute aqueous poly(vinyl alcohol) solutions. The range of sizes of the particles (fluorescent molecules, proteins, and polymers) was chosen to explore various length scales of the polymer solutions as defined by the polymer-polymer correlation length. For particles larger than the correlation length, we find that the diffusion coefficient, D, decreases exponentially with the polymer concentration. This can be explained by an exponential increase in the solution viscosity, consistent with the Stokes-Einstein equation. For probes on the order of the correlation length, the decrease of the diffusion coefficient cannot be accounted for by the Stokes-Einstein equation, but can be fit by a stretched exponential, D approximately exp(-alphacn), where we find n = 0.73-0.84 and alpha is related to the probe size. These results are in accord with a diffusion model of Langevin and Rondelez (Polymer 1978, 19, 1875), where these values of n indicate a good solvent quality.     

The rotational diffusion of chloroplast phosphate translocator and of lipid molecules in bilayer membranes

The rotational mobility of the phosphate translocator from the chloroplast envelope and of lipid molecules in the membrane of unilamellar azolectin liposomes has been investigated. The rotational dynamics of the liposome membrane were investigated by measuring the rotational diffusion of eosin-5-isothiocyanate(EITC)-labeled L-alpha-dipalmitoylglycerophosphoethanolamine (Pam2 GroPEtn) in the lipid phase of the vesicles, either in the presence or absence of the reconstituted phosphate translocator. The temperature dependence of the anisotropy decay showed that above 25 degrees C the main contribution to the anisotropy decay was caused by uniaxial anisotropic rotation of the labelled lipid molecules around the axis normal to the membrane plane. The rate of rotation of the labelled lipid molecules was strongly dependent on the viscosity of the medium (eta 1). Extrapolation to eta 1 = 0 Pa.s yielded a correlation time of phi = 20 +/- 5 ns, t = 30 degrees C, for lipid rotation with respect to the membrane normal. The rotational diffusion coefficient of the lipid molecules was calculated to be Dr = 2.0 x 10(9) rad2.s-1 and the apparent microviscosity in the vesicle membrane, as derived from the rotational correlation time, was eta 2 approximately 12 mPa.s. The rotational correlation time of the phosphate translocator in the membrane was only slightly dependent on the viscosity of the medium. The temperature dependence of the protein rotation also indicated that the rotation of the protein in the membrane was largely restricted and occurred mainly about the axis normal to the membrane plane. Measurements at a medium viscosity of eta 1 = 1 mPa.s yielded a value of phi r approximately 450 ns corresponding to Dr = 8.8 x 10(7) rad2.s-1 for protein rotation with respect to the membrane normal. From this value and the data of the lipid rotation, the cross-sectional area of the protein part embedded in the membrane was calculated to be approximately 9 nm2. This cross-sectional area is large enough to include at most 14 membrane-spanning helices. Our results also indicated that at lipid/protein molar ratios greater than or equal to 1.5 x 10(4): 1 aggregation occurred in the model membranes below 30 degrees C. However, above 30 degrees C and at a high dilution of the protein in the membrane it appeared that the membrane viscosity monitored by lipid and protein rotational diffusion were identical.     

Electric field-induced concentration gradients in planar supported bilayers.

A simple method of generating electric field-induced concentration gradients in planar supported bilayers has been developed. Gradients of charged, fluorescently labeled probes were visualized by epifluorescence microscopy and could be observed at field strengths as low as 1 V/cm. Steady-state concentration gradients can be described by a simple competition between random diffusion and electric field-induced drift. A model based on this principle has been used to determine the diffusion coefficient of the fluorescent probes. This technique achieves a degree of electrical manipulation of supported bilayers that offers a variety of possibilities for the development of new molecular architectures and the study of biological membranes.     

Chain stiffness of heteropolysaccharide from Aeromonas gum in dilute solution by dynamic light scattering

Dynamic light scattering measurements have been made on 15 fractions of aeromonas (A) gum, an extracellular heteropolysaccharide produced by the strain Aeromonas nichidenii, with dimethylsulfoxide containing 0.2M lithium chloride as the solvent at 25 degrees C. Data for the translational diffusion coefficient D covering a molecular weight range from 4.5 x 10(5) to 2.1 x 10(6) and ratios of the z-average radius of gyration (z) (1/2) to the hydrodynamic radius R(H) (calculated with previous (z) data) suggest that the polymer behaves like a semiflexible chain in this solvent similar to the stiffness of cellulose derivatives. Thus the D data are analyzed on the basis of the Yamakawa-Fujii theory for the translational friction coefficient of a wormlike cylinder by coarse-graining the heteropolysaccharide molecule. Excluded-volume effects are taken into account in the quasi-two-parameter scheme, as was done previously for (z) and [eta] (the intrinsic viscosity) of A gum in the same solvent. The molecular weight dependence of R(H) is found to be explained by the perturbed wormlike chain with a persistence length of 10 nm, a linear mass density of 1350 nm(-1), an excluded-volume strength parameter of 1.3 nm, and a chain diameter of 2.8 nm. These parameters are in substantial agreement with those estimated previously from (z) and [eta] data, demonstrating that the solution properties (D, (z), and [eta]) of the heteropolysaccharide are almost quantitatively described by the current theories for wormlike chains in the molecular weight range studied.     

Spectrofluorimetric methods for the measurement of the so called ”Microviscosity“ of lipidic structures

The cellular interactions involving the membrane depend on its physico-chemical nature and on the topographical distribution of the membrane receptors. At present, the role of the lipidic regions is not well defined; however, it is known that the fluidity or « microviscosityof the lipidic components controls important processes in cellular biology.Different spectrofluorimetric methods, continuous or time resolved, susceptible to the cohesion of lipidic regions have been developed:

1. The anisotropy of fluorescence where the rotation of probes is studied (linked to the coefficient of diffusional rotation).

2. The inhibition of fluorescence where the kinetics of the reaction is practically diffusion controlled.

3. The formation of emissive intramolecular complexes where the internal rotations of interchromophoric bonds are studied.

After the development of kinetic models, the methods have been tested with synthetic and natural organized assemblies.The values of « microviscosityobtained with these methods may be different because the environments of probes are different. Therefore, the concept of « microviscosity , applied to biological membranes is limited.     

Lateral diffusion on tubular membranes: quantification of measurements bias

Single Particle Tracking (SPT) is a powerful technique for the analysis of the lateral diffusion of the lipid and protein components of biological membranes. In neurons, SPT allows the study of the real-time dynamics of receptors for neurotransmitters that diffuse continuously in and out synapses. In the simplest case where the membrane is flat and is parallel to the focal plane of the microscope the analysis of diffusion from SPT data is relatively straightforward. However, in most biological samples the membranes are curved, which complicates analysis and may lead to erroneous conclusions as for the mode of lateral diffusion. Here we considered the case of lateral diffusion in tubular membranes, such as axons, dendrites or the neck of dendritic spines. Monte Carlo simulations allowed us to evaluate the error in diffusion coefficient (D) calculation if the curvature is not taken into account. The underestimation is determined by the diameter of the tubular surface, the frequency of image acquisition and the degree of mobility itself. We found that projected trajectories give estimates that are 25 to 50% lower than the real D in case of 2D-SPT over the tubular surface. The use of 3D-SPT improved the measurements if the frequency of image acquisition was fast enough in relation to the mobility of the molecules and the diameter of the tube. Nevertheless, the calculation of D from the components of displacements in the axis of the tubular structure gave accurate estimate of D, free of geometrical artefacts. We show the application of this approach to analyze the diffusion of a lipid on model tubular membranes and of a membrane-bound GFP on neurites from cultured rat hippocampal neurons.     

Dynamic imaging of lateral diffusion by electron spin resonance and study of rotational dynamics in model membranes. Effect of cholesterol.

The effects of cholesterol on the dynamics and the structural properties of two different spin probes, the sterol type CSL and the phospholipid type 16-PC, in POPC/cholesterol oriented multilayer model membranes were examined. Our results are consistent with a nonideal solution containing cholesterol-rich clusters created by the self association of cholesterol in POPC model membranes. The lateral diffusion coefficient D of the spin probes was measured over the temperature range of 15 to 60 degrees C and over the concentration range of 0 to 30 mol% of cholesterol in the model membrane by the electron spin resonance (ESR) imaging method. The rotational diffusion coefficients (including R perpendicular) and the order parameter S were determined utilizing a nonlinear least square ESR spectral simulation method. D, R perpendicular and S of CSL deviate considerably from linear dependence on mole percent cholesterol. The D of CSL was decreased by a factor of four at 15 degrees C and a factor of two at 60 degrees C for concentrations of cholesterol over 10 mol %, whereas those of 16-PC were hardly affected. Cholesterol decreased R perpendicular by a factor of 10 at 30 mol % of cholesterol, but it increased slightly that of 16-PC. A significant increase of S for CSL due to the presence of cholesterol was observed. It is shown how the difference in variation of S for CSL vs. 16-PC with composition may be interpreted in terms of their respective activity coefficients, and how a single universal linear relation is obtained for the S of both probes in terms of a scaled temperature. Simple but general correlations of D and of R perpendicular with S were also found, which aid in the interpretation of these diffusion coefficients.     

Determination of molecular weight and related hydrodynamic parameters of high molecular weight protein fraction from a few oilseeds

The hydrodynamic parameters of the major protein fraction, viz. arachin from groundnut, alpha-globulin from sesame seed, brassin (M) from mustard seed and helianthinin from sunflower seed, have been determined in a single solvent system (0.05 M Tris-HCl buffer, pH 7.5 containing 0.5 M sodium chloride): sedimentation coefficient (s0(20,w)) and diffusion coefficient (D0(20,w)) by analytical ultracentrifugation, intrinsic viscosity [eta] by Ostwald viscometry and partial specific volume (V) by densimetry. The molecular weights (M) of the four proteins, calculated using the sedimentation-viscosity and sedimentation-diffusion coefficient methods, were found close to each other. The values have been compared with those in the literature and the reasons for discrepancies have been discussed.     

Order and fluidity of lipid membranes as determined by fluorescence anisotropy decay

The fluorescence anisotropy decay of four different probes in bilayers of dimyristoylphosphatidylcholine was measured. The probes are diphenylhexatriene, diphenyloctatetraene, trimethylaminodiphenylhexatriene, and trans-parinaric acid. The data for each probe were analyzed in terms of two orientational order parameters, the ordinary order parameter and a higher one, and two rotational diffusion coefficients. The order parameters are largely independent of probe size, but depend on the position of the probes along the membrane normal, thus reflecting the profile of lipid order. If a probe is located in the plateau region of lipid order, its order parameters are interpreted as representing the rigid-body order of lipids. According to this interpretation, the total lipid order in the plateau region originates about equally from rigid-body order and conformational order. The two order parameters obtained for each probe are used to derive approximate angular distributions of the probe molecules. The diffusion coefficient for rotation about the long molecular axis is found to be infinitely large, indicating unhindered rotation about this axis. The diffusion coefficient for rotation about the short molecular axes is evaluated for a viscosity which results as 0.2 poise. This viscosity for rotational diffusion is an order of magnitude smaller than the viscosity for lateral diffusion indicating that at least two viscosities are required to characterize the fluidity of a lipid membrane.Abbreviations FAD fluorescence anisotropy decay - DMR deuterium magnetic resonance - ESR electron spin resonance - DMPC dimyristoylphosphatidylcholine - DPPC dipalmitoylphosphatidylcholine - DPH 1,6-diphenyl-1,3,5-hexatriene - DPO 1,6-diphenyl-1,3,5,7-octatetraene - TMA-DPH 1-[4-(trimethylamino)phenyl]-6-phenyl-1,3,5-hexatriene - tPnA trans-parinaric acid - NPN N-phenyl-1-naphthylamine - BBO 2,5-bis(4-biphenylyl)oxazole     

The lack of relationship between fluorescence polarization and lateral diffusion in biological membranes

An investigation has been carried out of the relationship between changes in the fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) and concomittant changes in the lateral diffusion of proteins and lipid probes in membranes. Plasma membranes from lymphocytes and a CH1 mouse lymphoma line were treated with up to 70 mol% (relative to the total membrane phospholipid) of oleic or linoleic fatty acids. Under these conditions the fluorescence polarization of DPH decreased by between 8 and 15% which, in the framework of the microviscosity approach, suggests a membrane fluidity change of between 20 and 50%. The lateral diffusion coefficients of surface immunoglobin and the lipid probes 3,3′-dioctadecylindocarbocyanine and pyrene were also measured in these membranes using the fluorescence photobleaching recovery technique and the rate of pyrene excimer formation. The diffusion rates were found to be unaffected by the presence of free fatty acids. Hence despite large ‘microviscosity’ changes as reported by depolarization of DPH fluorescence, lateral diffusion coefficients are essentially unchanged. This finding is consistent with the idea that perturbing agents such as free fatty acids do not cause a general fluidization of the membrane but act locally to alter, for example, protein function. It is also consistent with the suggestion that lateral mobility of membrane proteins is not modulated by the lipid viscosity.     

A milling crowd model for local and long-range obstructed lateral diffusion. Mobility of excimeric probes in the membrane of intact erythrocytes.

A new model for lateral diffusion, the milling crowd model (MC), is proposed and is used to derive the dependence of the monomeric and excimeric fluorescence yields of excimeric membrane probes on their concentration. According to the MC model, probes migrate by performing spatial exchanges with a randomly chosen nearest neighbor (lipid or probe). Only nearest neighbor probes, one of which is in the excited state, may form an excimer. The exchange frequency, and hence the local lateral diffusion coefficient, may then be determined from experiment with the aid of computer simulation of the excimer formation kinetics. The same model is also used to study the long-range lateral diffusion coefficient of probes in the presence of obstacles (e.g., membrane proteins). The dependence of the monomeric and excimeric fluorescence yields of 1-pyrene-dodecanoic acid probes on their concentration in the membranes of intact erythrocytes was measured and compared with the prediction of the MC model. The analysis yields an excimer formation rate for nearest neighbor molecules of approximately 1 X 10(7) s-1 and an exchange frequency of approximately greater than 2 X 10(7) s-1, corresponding to a local diffusion coefficient of greater than 3 X 10(-8) cm2 s-1. This value is several times larger than the long-range diffusion coefficient for a similar system measured in fluorescence photobleaching recovery experiments. The difference is explained by the fact that long-range diffusion is obstructed by dispersed membrane proteins and is therefore greatly reduced when compared to free diffusion. The dependence of the diffusion coefficient on the fractional area covered by obstacles and on their size is derived from MC simulations and is compared to those of other theories lateral diffusibility.     

Transmembrane Protein Diffusion in Gel-Supported Dual-Leaflet Membranes

Tools to measure transmembrane-protein diffusion in lipid bilayer membranes have advanced in recent decades, providing a need for predictive theoretical models that account for interleaflet leaflet friction on tracer mobility. Here we address the fully three-dimensional flows driven by a (nonprotruding) transmembrane protein embedded in a dual-leaflet membrane that is supported above and below by soft porous supports (e.g., hydrogel or extracellular matrix), each of which has a prescribed permeability and solvent viscosity. For asymmetric configurations, i.e., supports with contrasting permeability, as realized for cells in contact with hydrogel scaffolds or culture media, the diffusion coefficient can reflect interleaflet friction. Reasonable approximations, for sufficiently large tracers on low-permeability supports, are furnished by a recent phenomenological theory from the literature. Interpreting literature data, albeit for hard-supported membranes, provides a theoretical basis for the phenomenological Stokes drag law as well as strengthening assertions that nonhydrodynamic interactions are important in supported bilayer systems, possibly leading to overestimates of the membrane/leaflet viscosity. Our theory provides a theoretical foundation for future experimental studies of tracer diffusion in gel-supported membranes.     

Nanosecond fluorescence anisotropy decays of 1,6-diphenyl-1,3,5-hexatriene in membranes.

Nanosecond decays of the fluorescence anisotropy, r, were studied for the emission of 1,6-diphenyl-1,3,5-hexatriene (DPH) embedded in a series of mixed multilamellar liposomes containing egg yolk phosphatidylcholine, phosphatidylethanolamine and cholesterol in varying molar ratios, as well as in membranes of intact cells and in virus envelopes. The relative contributions of the fast and the infinitely slow decaying component to the steady-state value r, of the fluorescence anisotropy were very similar for artifical and biological membranes. Angles, theta, of the cone, by which the motion of the fluorescent molecule is limited, were calculated from the intensity of the infinitely slow decaying anisotropy component and compared with steady-state fluorescence anisotropies and with 'microviscosities', (eta). An increase in (eta) from 1.5 to 5.2 P in our systems was accompanied by a decrease in theta from 49 degrees to 30 degrees while the decrease in the mean motional relaxation times, phi f, of the label molecule was not more than 1 ns and due mainly to changes in the potential, by which the diffusion of DPH in the membrane is restricted. From these observations we conclude that differences in the steady-state fluorescence anisotropy and in 'microviscosities' of cholesterol-containing membranes (r greater than 0.15) represent changes in the degree of static orientational constraint rather than changes in diffusion rates of the label.     

Lateral diffusion in substrate-supported lipid monolayers as a function of ambient relative humidity

We analyzed the influence of water activity on the lateral self-diffusion of supported phospholipid monolayers. Lipid monolayer membranes were supported by polysaccharide cushions (chitosan and agarose), or glass. A simple diffusion model was derived, based on activated diffusion with an activation energy, E(a), which depends on the hydration state of the lipid headgroup. A crucial assumption of the derived model is that E(a) can be calculated assuming an exponential decay of the humidity-dependent disjoining pressure in the monolayer/substrate interface with respect to the equilibrium separation distance. A plot of ln(D) against ln(p(0)/p), where D is the measured diffusion coefficient and p(0) and p are the partial water pressures at saturation and at a particular relative humidity, respectively, was observed to be linear in all cases (i.e., for differing lipids, lateral monolayer pressures, temperatures, and substrates), in accordance with the above-mentioned diffusion model. No indications for humidity-induced first-order phase transitions in the supported phospholipid monolayers were found. Many biological processes such as vesicle fusion and recognition processes involve dehydration/hydration cycles, and it can be expected that the water activity significantly affects the kinetics of these processes in a manner similar to that examined in the present work.     

Effect of Geometrical and Chemical Constraints on Water Flux across Artificial Membranes

Studies have been made on the temperature dependence of both the hydraulic conductivity, L_p, and the THO diffusion coefficient, ω, for a series of cellulose acetate membranes (CA) of varying porosity. A similar study was also made of a much less polar cellulose triacetate membrane (CTA). The apparent activation energies, E_a, for diffusion across CA membranes vary with porosity, being 7.8 kcal/mole for the nonporous membrane and 5.5 kcal/mole for the most porous one. E_a for diffusion across the less polar CTA membrane is smaller than E_a for the CA membrane of equivalent porosity. Classical viscous flow, in which the hydraulic conductivity is inversely related to bulk water viscosity, has been demonstrated across membranes with very small equivalent pores. Water-membrane interactions, which depend upon both chemical and geometrical factors are of particular importance in diffusion. The implication of these findings for the interpretation of water permeability experiments across biological membranes is discussed.     

Red blood cell orientation in orbit C = 0.

Two modes of behavior of single human red cells in a shear field have been described. It is known that in low viscosity media and at shear rates less than 20 s-1, the cells rotate with a periodically varying angular velocity, in accord with the theory of Jeffery (1922) for oblate spheroids. In media of viscosity greater than approximately 5 mPa s and sufficiently high shear rates, the cells align themselves at a constant angle to the direction of flow with the membrane undergoing tank-tread motion. Also, in low viscosity media, as the shear rate is increased, more and more cells lie in the plane of shear, undergoing spin with their axes of symmetry aligned with the vorticity axis of the shear field in an orbit "C = 0" (Goldsmith and Marlow, 1972). We have explored this latter phenomenon using two experimental methods. First, the erythrocytes were observed in the rheoscope and their diameters measured. Forward light scattering patterns were correlated with the red cell orientation mode. Light flux variations after flow onset or stop were measured, and the characteristic times of erythrocyte orientation and disorientation were assessed. The characteristic time of erythrocyte orientation in Orbit C = 0 is proportional to the inverse of the shear rate. The corresponding coefficient of proportionality depends on the suspending medium viscosity eta o. The disorientation time tau D, after flow has been stopped, is such that the ratio tau D/eta o is independent of the initial applied shear stress. However, tau D is much shorter than one would expect if pure Brownian motion were involved. The proportion of erythrocytes in orbit C = 0 was also measured. It was found that this proportion is a function of both the shear rate and eta o. At low values of eta o, the proportion increases with increasing shear rate and then reaches a plateau. For higher values of eta o (5 to 10 mPa s), the proportion of RBC in orbit C = 0 is a decreasing function of the shear stress. A critical transition between orbit C = 0 and parallel alignment was observed at high values of eta o, when the shear stress is on the order of 1 N/m2. Finally, the effect of altering membrane viscoelastic properties (by heat or diamide treatment) was tested. The proportion of oriented cells is a steep decreasing function of red cell rigidity.     

Hydrodynamic properties of human cervical-mucus glycoproteins in 6M-guanidinium chloride.

Cervical mucins and fragments thereof were studied by sedimentation-velocity, rotatory viscometry and laser light-scattering performed as photon-correlation spectroscopy as well as low-angle total-intensity measurements. The Mr of the whole mucins is 10 X 10(6)-15 X 10(6), whereas fragments obtained after reduction of disulphide bonds ('subunits') have Mr 2.1 X 10(6)-2.9 X 10(6), depending on the method used. Subsequent trypsin digestion of subunits afforded glycopeptides with Mr approx. 0.4 X 10(6). The high frictional ratio for the whole mucins is interpreted as a large degree of expansion. The Stokes radius calculated from the diffusion coefficient is approx. 110nm for the whole mucins, which is in agreement with that estimated from the radius of gyration (130nm) by using the concept of the equivalent hydrodynamic sphere. The ratio of the concentration-dependence parameter for the reciprocal sedimentation coefficient (Ks) to the intrinsic viscosity ( [eta] ) for the whole mucins is 1.42, suggesting that the individual macromolecule occupies a spheroidal domain in solution. The relationship between [eta] and Mr for whole mucins, subunits and T-domains suggests that they are linear flexible macromolecules behaving as somewhat 'stiff' random coils. This conclusion is supported by the relationships between the sedimentation coefficients, the diffusion coefficients and the Mr. The hydrodynamic behaviour of the mucins is thus close to that expected for coiling macromolecules entrapping a lot of solvent, which is consistent with the postulated polymeric structure.