Coupled size-exclusion columns

A set of bimodal columns is useful in this case. These consist of two or more columns with two specific pore sizes which differ by a factor of 10 (e.g. 100 and 1000 A columns). If the pore volume of both gel types is the same, then a calibration graph with optimum linearity is obtained. 

Note that a sample component which migrates nonselectively through a column, i.e. is subject to total exclusion or total permeation, inevitably undergoes unnecessary band broadening and hence impairs the resolution. The mobile phase should pass through the columns according to increasing pore diameter. 

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Chromatographic approaches have been also used to separate nanoparticles from samples coupled to different detectors, such as ICP-MS, MS, DLS. The best known technique for size separation is size exclusion chromatography (SEC). A size exclusion column is packed with porous beads, as the stationary phase, which retain particles, depending on their size and shape. This method has been applied to the size characterization of quantum dots, single-walled carbon nanotubes, and polystyrene nanoparticles [168, 169]. Another approach is hydro-dynamic chromatography (HDC), which separates particles based on their hydro-dynamic radius. HDC has been connected to the most common UV-Vis detector for the size characterization of nanoparticles, colloidal suspensions, and biomolecules [170-172]. 

Gardner et al. [9] recognised that because component separations on size exclusion columns with distilled water are affected by chemical physical interactions as well as component molecular size, distilled water size exclusion chromatography will also fractionate dissolved metal forms. These workers interfaced distilled water size exclusion chromatography with inductively coupled argon plasma detection to fractionate and detect dissolved forms of calcium and magnesium in lake and river waters. 

Nerfin, C. Salafranca, J. Cacho, J. Rubio, C. Separation of polymer and on-line determination of several antioxidants and UV stabilizers by coupling size-exclusion and normal-phase high-performance liquid chromatography columns. J. Chromatogr., A 1995, 690, 230-236. 

The first organometallic miktoarm star copolymer, PFS(PI)3, with PDI of 1.04 was synthesized through an anionic polymerization by using SiCl4 as a coupling agent, as shown in Scheme 3.11.32 The well-defined structure was confirmed by the characterizations of GPC and H nuclear magnetic resonance (NMR) spectroscopy. The PFS(PI)3 miktoarm star copolymer was obtained in a moderate yield after size-exclusion column purification (Mn = 21,300 PDI = 1.05) and with a composition ratio of PFS PI = 1 9.5. 

This method employs SEC coupled to electro-spray ionization mass spectrometry (ESI/MS) and can he applied to culture filtrate samples and crude PIA extracts isolated from cell surfaces. The acquisition of ESI mass spectra of PIA in positive ion mode is based on the ionization of groups such as hydroxyl groups on sugar residues, or in the case of deacetylated PIA on already ionized amino groups. On the size exclusion columns used under the specific buffer conditions, PIA elutes earlier and separate from most other molecules, soon after the exclusion volume. 

As with other size-exclusion techniques, the pore size of the selected Zorbax GF column should provide resolution over the molecular size range of the proteins that are to be separated. The Zorbax GF-250 column separates proteins in the range of 4000 to 400,000 Da. The Zorbax GF-450 provides separation over the range of 10,000 to 1,000,000 Da. When these two columns are coupled, they can be used to separate proteins with molecular weights of 4000 to 1,000,000. 

Stopher and Gage used size-exclusion chromatography (SEL) coupled to reversed phase HPLC for the direct injection of plasma in the analysis of an antifungal agent, voriconazole (12). Their system consisted of three columns, i.e. first a size-exclusion... 

Advances in size-exclusion chromatography, coupled with refractive index, absorption, viscosity, and lightscattering detectors, and MALDI-ToFMS, have made it possible to accurately determine molecular weight distribution (oligomer profiling), even at the relatively low values of polymeric additives (up to about 5000 Da). Advances in column design, e.g. high-resolution PS/DVB columns (> 105 plates m-1) mean that SEC can provide a valuable alternative to conventional HPLC techniques for the separation of small molecules. 

Polymer gel GPC columns packed with 10-pm gels can exhibit efficiencies of 12,000 to 16,000 plates depending on the pore size. Single columns of this type produce bandwidths from 160 to 180 pi. As columns are coupled in series, bandwidth increases as the square root of the number of columns, as may be seen from Equation 1. Plate number doubles, but so does the exclusion volume. The 5-pm gel columns typically achieve 20,000 to 24,000 plates, and are represented by the bottom two rows of the table. The implications of the bandwidth values in Table I will be discussed below. 

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