Particle Centrifugation Electrophoresis

Differences in mobilities of ions, molecules, or particles in an electric field can be exploited to perform useful separations. Primary emphasis is placed on electrophoresis and dielec trophoresis. Analogous separation processes involving magnetic and centrifugal force fields are widely apphed in the process industiy (see Secs. 18 and 19). 

The opposite effect to electrophoresis is the generation of a sedimentation potential. If a charged particle moves in the gravitational field or in a centrifuge, an electric potential arises — the sedimentation potential. While the particle moves, the ions in the electric double layer lag somewhat behind due to the liquid flow. A dipole moment is generated. The sum of all dipoles causes the sedimentation potential. 

We should note that almost every system, even those in the c class, contains a discontinuity in pi located at the walls, electrodes, and other nonpermeable barriers that define the physical limits of the system. However, although walls represent an abrupt pi barrier, we do not automatically put separation in the d class because of ttfelm. If they are passive container walls simply holding the system in place, or if they are electrodes which charged particles never touch (as in electrophoresis), a d classification is not justified. However, if the walls play a major part in structuring the separation (such as membrane filters or the outer wall of a centrifuge cell which structures the solute concentration profiles in equilibrium sedimentation), those walls convey a d classification to the method. 

A relatively simple and quick procedure for the isolation of Photosystem I-enriched particles from the thermophilic cyanobacterium Phormidium laminosum, without the use of detergents for solubilization, is described. The procedure involves sonication of cells, centrifugation and DEAE-cellulose chromatography. The particles had an 02 uptake activity of up to 200 pmol 02. mg chlorophyll h 1 and appeared as vesicles of 200 100 nm diameter when observed under electron microscopy. The analysis of the chlorophyll-protein complexes by polyacrylamide gel electrophoresis showed that these particles are enriched in the complexes associated with Photosystem I and partially depleted in those associated with Photosystem II. The particles did not contain ferredoxin and were active in NADP-photoreduction only in the presence of added ferredox in. They were also able to photoreduce externally added electron mediators using ascorbate as electron donor, the reduced mediators can be coupled to hydrogenase for the production of H2 or for the activation of cyanobacterial phosphoribulokinase using a ferredoxin/thioredoxin system. 

Centrifugally accelerated electrophoresis, as described by McDonald et al. [26], is based on the fact that centrifugal force drifts the electrolyte (mobile phase) from the centre of the circular paper sheet to the collecting cups, and the electric field that acts perpendicularly to the direction of the centrifugal force drifts the charged particles to the positive and negative poles. 

Emphasis is on rational theory and its consequences, with the purpose of showing the underlying unity of PCH, in which diverse phenomena can be described in physically and mathematically similar ways. The magic of this unity is shown in the similar manner in which solutes concentrate in a flow containing chemically reacting surfaces, reverse osmosis membranes, and electrodialysis membranes or the similarity of particle motions in sedimentation, centrifugation, ultrafiltration, and electrophoresis. Experimental results, numerical solutions, and reference to topics not covered are noted where they serve to illustrate a concept, result, or limitation of what has been presented. Empiricism is not eschewed, but only limited use is made of it and then only when it contributes to a better understanding of an idea or phenomenon. 

Unfortunately, it is not yet possible to purify informosomes free of contaminating ribosomes, subribosomal particles, and soluble proteins of the cytoplasm. The only useful purification method is centrifugation in a CsCl density gradient, but this requires a prefixation with formaldehyde or with other cross-linking agents (Spirin et al., 1965). The fixation makes it impossible to study adequately the proteins of particles or their activities. Another possible approach is to use free flow electrophoresis, which should differentiate ribosomes and D-RNP, but such work has only just started (Schweiger and Hannig, 1970). 

Frictional coefficient A coefficient that determines the frictional force on a particular particle (such as a molecule) in a particular medium at a given velocity. In the context of electrophoresis or centrifugation, it determines how fast a chemical species will move in a particular medium in response to a given electrical field or centrifugal force. 

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