Ultracentrifuge measurements

In a centrifugal field, dissolved molecules or suspended particles either sediment (if their density exceeds that of the pure solvent), or flotate for the opposite case (negative or inverse sedimentation). Under otherwise identical experimental conditions, the velocity of the molecules or particles depends on the viscosity of the solution or suspension and - very importantly - on the mass and shape of the dissolved species. Sedimentation and flotation are antagonized by the diffusion. Depending on the rotor speed and the molar mass of the dissolved/dis- 

The proportionality factor F is called friction factor and is identical for diffusion and sedimentation. Using the Einstein Sutherland equation 

If (1) the diffusion coefficient D of the polymer in the used solvent, (2) the specific volume v of the dissolved polymer, and (3) the density of the solvent solvent known, one can determine the molecular weight of the dissolved polymer according to the above equation by measuring the sedimentation coefficient (by measuring the maximum of the concentration gradient at regular time intervals). 

The thus determined molecular weight is an apparent one since s and D depend on the polymer concentration. Therefore, extrapolation to concentration zero is required. The sedimentation coefficient obtained by extrapolating c — 0 is called sedimentation constant Sn  

Here is a constant which depends on solvent and temperature. Thus the sedimentation constant can be calculated from they-axis intercept when the reciprocal of the sedimentation coefficient s determined at different polymer con- 

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Paudjojo was able to show by ultracentrifuge measurements that there is a strong specific interaction between (Lys) and CIO4, however no measurable one between (Lys)n and SO According to her results, the sedimentation coefficient... 

Solvent extraction measurements and ultracentrifuge measurements have been reported only in single instances but agreement with other methods has been observed for U0 (N0 ) (Goldberg,... 

The fi isomer of [(B W Cy]6 ( tungstate X ) is difficult to isolate in pure form, but can be obtained indirectly from reduced tp. There seems little doubt, based on comparisons with heteropolyanions, and from JH and 183W NMR, that the anion structure corresponds to that of /S-[SiWi2O40]4-.52 The composition and structure of tp are still unknown. Although W6 and W24 formulas have been suggested, recent ultracentrifugation measurements indicate a dodecamer. Reduction of tp by two equivalents/12W causes transformation to a new species tp, which, upon reoxidation, slowly is converted to tp. potassium salt of oxidized tp has been isolated. 

Ultracentrifuge measurements indicate that the glycogen molecule in solution is not spherical, but possesses an elongated form. 

Other Physical Measurements. Viscosity measurements did not show differences between samples treated with acetyl glutamate and controls, before and after storage in the cold (35). Extensive series of ultracentrifuge measurements did not clarify the phenomenon the latter studies were conducted at several protein and acetyl glutamate concentrations, and at temperatures varying from 5° to 20° only after cold inactivation for 40 hours were marked changes noticeable. 

Although all ultracentrifugation measurements were done in buffers which contained no excess zinc, bound zinc ions are required for the specific homodimer formation of ZDD as the apo form of the domain was shown to be unfolded and nonspecifically aggregated [5]. Atomic absorption spectroscopy of the samples in the buffers used for these experiments confirmed the expected stoichiometry of zincbinding. 

This ol)servation appears to have been overlooked by many workers. A few results reported since 1936 are summarized in Table I. Essentially similar results have been obtained by ultracentrifuge measurements on 20 samples of glycogen, all of which were polymolecular the molecular weights varied between (2-6) X 10, six of the preparations being polydisperse. Polglase and coworkers Ukewise found samples of human glycogen to be... 

Ultracentrifuge measurements (113, 128) indicate that 6-tungstoselenate(IV) is a tetramer, Se4W24096Hj. The apparent formula weight of the sodium salt, 6450, is the second-highest ever reported for a heteropoly compound. Confirmation of this would be most desirable. 

Using the ultracentrifuge, measurements have been made on the... 

On the addition of oxalate, blood separates into two layers. A dark red layer contains the blood corpuscles and a yellowish layer contains the plasma and plasma proteins. The plasma proteins can then be separated by fractional precipitation with ammonium sulfate solution. A 20%-25% ammonium sulfate solution will precipitate fibrinogens a 33% solution, globulin and a 50% solution, pseudoglobulin while albumins only precipitate at very high ammonium sulfate concentration. Ultracentrifuge measurements enable four components (X, A, G, M) with different sedimentation coefficients to be differentiated and isolated ... 

Light scattering Ultracentrifugation Measurement of the intensity of light scattered by dilute polymer solution. The intensity is measured as a function of concentration and angle of observation Ww Hundreds to tens of millions... 

Fig.3. Sedimentation pattern of normal human serum in the ultracentrifuge, measured with a schlieren optical system. Left hand picture taken 51 min after start of centrifugation right hand picture after 125 min. Centrifugation at 59,800 rpm. A albumin, 4.5S G = globulin, 7S M = macroglobulin, 19S.

Determination of molecular symmetry. Molecular symmetry of P. can be determined from measurements of viscosity, streaming birefringence, rates of sedimentation and diffusion, or directly by electron microscopy, For a known M, the frictional coefficient can be calcnlated from ultracentrifugal measurements, e.g. from the sedimentation /= [Mfl - vp)]/S, where v is the partial specific volume, p is the density and S is the sedimentation constant. The axial ratio a/b of a P. can be derived from the frictional ratiowhereis the/of a spherical molecule. The value of a/b for most globular P. is between 2 and 20, and greater than 20 for fibrous R, e, g. the axial ratio of fibrinogen is 30. 

The mucin precipitated from synovial fluids by weak acids appears to be a dissociable complex of hyaluronic acid and proteins. Electrophoretic studies of synovial fluids (at pH 8.6) have usually shown the presence of one fast-moving component considered to be free hyaluronic acid, but some fluids show a second fast component, possibly a hyaluronic acid -- protein complex (69). On the basis of studies in the ultracentrifuge, the mucin possibly may exist in synovial fluid partially as a mucoprotein with a molecular weight of one to ten million (70). Electrophoretic and ultracentrifugal measurements at varying pH values provide two of the best ways of demonstrating interaction in such systems. 

It is now apparent from electrophoresis and ultracentrifuge measurements, correlated with separation by neutral salts (167, 208, 209, 210, 211), that the latter are incapable by any known means of accurately fractionating protein mixtures without complicating the operations to a point at which they are of little use as practical analytical procedures. Thus, Dole (212) found that the albumin-globulin ratio measured electrophoretically was about 2/3 of the ratio fotmd by chemical fractionation, an observation which was also checked by Pillemer and Hutchinson (196) by use of a methanol fractionation which was checked against electrophoresis. 

Similar experiments aimed to refold the large subunits, showed that the absorption spectrum does not completely return to the initial value after incubation, the reversibility being of the order of 50%. Besides ultracentrifuge measurements showed that, under these con-... 

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