Sedimentation-equilibrium technique

Sedimentation equilibrium technique, 19 256 Sedimentation velocity technique, 19 256 Sediments, arsenic in marine samples, 44 149, 162-164, 169, 181 [Se U ] cations, 35 297-298 Selective vaporization, for preparation of actinide metals, 31 12-13, 26 Selenide, production, 38 82 Selenium... 

Molecular weight (MW) of the purified enzyme in 0.1 M NaCl, 0.01 M sodium phosphate buffer, pH 7 (NaCl-NaP04 buffer), has been established by both low- and high-speed sedimentation equilibrium techniques (13, 16, 17). 

The disadvantage of the sedimentation equilibrium technique is that the establishment of equilibrium may take as long as several days, which not only is inconvenient generally, but also accentuates the importance of avoiding convectional disturbances. 

The mass of a protein can be directly determined by sedimentation equilibrium, in which a sample is centrifuged at relatively low speed so that sedimentation is counterbalanced by diffusion. The sedimentation-equilibrium technique for determining mass is very accurate and can be applied under nondenaturing conditions in which the native quaternary structure of multimeric proteins is preserved. In contrast, SDS-polyacrylamide gel electrophoresis (Section 4.1.4) provides an estimate of the mass of dissociated polypeptide chains under denaturing conditions. Note that, if we know the mass of the dissociated components of a multimeric protein as determined by SDS-polyacrylamide analysis and the mass of the intact multimeric protein as determined by sedimentation equilibrium analysis, we can determine how many copies of each polypeptide chain is present in the multimeric protein. 

The mass of a protein can be directly determined by sedimentation equilibrium, in which a sample is centrifuged at relatively low speed so that sedimentation is counterbalanced by diffusion. The sedimentation-equilibrium technique for determining mass is very accurate and can be applied without denaturing the protein. Thus the native quaternary structure of multimeric proteins is preserved In contrast, SDS-polyacrylamide gel electrophoresis... 

The sedimentation equilibrium technique yields directly the particle mass M from the concentration profile along the radial distance r from the center of rotation ... 

The pronounced sensitivity of the light-scattering method to impurities of high molecular weight makes the method less attractive for measuring the molecular weights of hardwood xylans, which have relatively short molecules. The sedimentation-equilibrium technique appears more promising in this respect, especially since the time to reach equilibrium can be much... 

In the sedimentation equilibrium technique the centrifuge is operated at a lower speed—e.g., about 8000 rpm for a molecular weight of 60 000. When the centrifuge... 

Initially this method had the advantage of rapidity over the conventional sedimentation equflibtliun method. However, since it d iends upon the data from the extremities of the solution colunm, precise measurements are not possible and the method is inferior in accuracy. Moreover, since the revival of sedimentation equilibrium technique with specialty designed small and narrow cells and better suited (pities (Rayleigh interfermce), this method has mme or less fallen into disuse. 

At first glance, the contents of Chap. 9 read like a catchall for unrelated topics. In it we examine the intrinsic viscosity of polymer solutions, the diffusion coefficient, the sedimentation coefficient, sedimentation equilibrium, and gel permeation chromatography. While all of these techniques can be related in one way or another to the molecular weight of the polymer, the more fundamental unifying principle which connects these topics is their common dependence on the spatial extension of the molecules. The radius of gyration is the parameter of interest in this context, and the intrinsic viscosity in particular can be interpreted to give a value for this important quantity. The experimental techniques discussed in Chap. 9 have been used extensively in the study of biopolymers. 

SEC-MALLS and SEC-LALLS Coupled chromatography and light-scattering photometry that allows the determination of a number of important values along with chain length distribution, sedimentation equilibrium experiment Ultracentrifugation technique that allows chain length information to be determined. 

It might be noted that sedimentation equilibrium is approached very slowly however, techniques that permit equilibrium conditions to be estimated from preequilibrium measurements have been developed by W. J. Archibald. Equations (86) and (87) predict a linear semilogarithmic plot of c versus x or x2 for gravitational and centrifugal studies, respectively. The slope of such a plot is proportional to the mass of the particles involved. Remember that monodispersity was assumed in the derivation of these equations. If this condition is not met for an experimental system, the plot just described will not be linear. If each particle size present is at equilibrium, however, each component will follow the equations and the experimental plot will be the summation of several straight lines. Under certain conditions these may be resolved to give information about the polydispersity of the system. In any event, nonlinearity implies polydispersity once true equilibrium is reached. 

The analysis of mixed associations by light scattering and sedimentation equilibrium experiments has been restricted so far to ideal, dilute solutions. Also it has been necessary to assume that the refractive index increments as well as the partial specific volumes of the associating species are equal. These two restrictions are removed in this study. Using some simple assumptions, methods are reported for the analysis of ideal or nonideal mixed associations by either experimental technique. The advantages and disadvantages of these two techniques for studying mixed associations are discussed. The application of these methods to various types of mixed associations is presented. 

We have shown some of the ways that mixed associations could be analyzed by light scattering, Archibald, or sedimentation equilibrium experiments. It might be appropriate at this point to make a comparison of some advantages and disadvantages of each technique. 

From a theoretical analysis of the intermediate stages during which the solute is being redistributed, Archibald136 has developed a technique which involves measurements at intervals during the early stages of the sedimentation equilibrium experiment and so does not entail a long wait for equilibrium to be established. The ratio s/D can be calculated from the expression... 

As stated in Section 8.2.1, only limited results concerning Mw determinations of lignins with the use of LALLS photometry have been published. This radically new and sophisticated technique constitutes an improvement over other methods, such as sedimentation equilibrium and size exclusion chromatography, for the evaluation of Mw. Its low angle capability, combined with high sensitivity, small sample size, and simplified clarification procedures, allow simple and accurate determinations to be made even on lignin samples which have been considered very troublesome. Moreover, as stated in Section 8.2.2, additional difficulties, inherent lignin properties, i.e., absorbance, fluorescence and anisotropy, may also be overcome. 

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