Separation techniques size exclusion chromatography

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]. 

Inductively coupled plasma-mass spectrometry (ICP-MS) is a powerful technique that uses an inductively coupled plasma as an ion source and a mass spectrometer as an ion analyzer. It can measure the presence of more than 75 elements in a single scan, and can achieve detection limits down to parts per trillion (ppt) levels for many elements—levels that are two or three orders of magnitude lower than those obtained by ICP-AES (Keeler 1991). It is more expensive than ICP-AES and requires more highly skilled technical operation. Aluminum levels in urine and saliva were detected down to 0.02 g/mL and in blood serum to 0.001 g/mL using ICP-MS (Ward 1989). Speciation studies have employed ICP-MS as a detector for aluminum in tissue fractions separated by size-exclusion chromatography (SEC) with detection limits of 0.04 g/g in femur, kidney and brain (Owen et al. 1994). 

The size of impurities such as fullerenes, metal nanoparticles, and polyaromatic carbons are much smaller than the carbon nanotubes. Such impurities can be separated using size exclusion chromatography (SEC). The carbon nanotubes are first dispersed in organic solvent either by functionalization or using smfactants. The dispersed SWNTs in organic solvents at low concentration can be used to separate the impurities by SEC techniques. The conventional styrene-... 

Names such as gel filtration chromatography (mobile phase is water), used by biochemists, and gel permeation chromatography (mobile phase is an organic solvent), used by polymer chemists, describe this technique. Size exclusion chromatography, however, is the recommended term. Molecular weight distribution of polymers can be obtained by this technique, and proteins, enzymes, peptides, nucleic acids, hormones, polysaccharides, and so on can be separated. 

Molecular weight distribution information obtained by size-exclusion chromatography on its own is insufficient to characterize the properties of complex polymers, such as copolymers and block and graft polymers [23,514,524]. For these polymers the chemical composition and functionality type distributions are equally important. A major obstacle to the characterization of these materials is that their molecular properties are present as joint distributions. Unlike the mass distribution the composition and functionality distributions can only be determined by separation methods that employ interactions with the stationary phase. To fully characterize a complex polymer it is not unusual to use manual or automated tandem techniques where the sample is fractionated according to its chemical or end group composition for subsequent further separation by size-exclusion chromatography to establish their mass distribution. Chromatographic methods may also be combined with spectroscopic methods to determine microstructural information. 

Proteins can be separated by any of a number of different HPLC techniques size-exclusion chromatography, ion-exchange chromatography, hydrophobic interaction chromatography, or reversed-phase chromatography. Affinity chromatography is not usu y considered to be an HPLC technique, and is not covered in this book. 

A number of analytical techniques such as FTIR spectroscopy,65-66 13C NMR,67,68 solid-state 13 C NMR,69 GPC or size exclusion chromatography (SEC),67-72 HPLC,73 mass spectrometric analysis,74 differential scanning calorimetry (DSC),67 75 76 and dynamic mechanical analysis (DMA)77 78 have been utilized to characterize resole syntheses and crosslinking reactions. Packed-column supercritical fluid chromatography with a negative-ion atmospheric pressure chemical ionization mass spectrometric detector has also been used to separate and characterize resoles resins.79 This section provides some examples of how these techniques are used in practical applications. 

In addition to the insoluble polymers described above, soluble polymers, such as non-cross-linked PS and PEG have proven useful for synthetic applications. However, since synthesis on soluble supports is more difficult to automate, these polymers are not used as extensively as insoluble beads. Soluble polymers offer most of the advantages of both homogeneous-phase chemistry (lack of diffusion phenomena and easy monitoring) and solid-phase techniques (use of excess reagents and ease of isolation and purification of products). Separation of the functionalized matrix is achieved by either precipitation (solvent or heat), membrane filtration, or size-exclusion chromatography [98,99]. 

The two techniques differ in that HDC employs a nonporous stationary phase. Separation is affected as a result of particles of different size sampling different velocities in the interstitial spaces. Size exclusion chromatography is accomplished by superimposing a steric selection mechanism which results from the use of a porous bed. The pore sizes may vary over a wide range and the separation occurs as a result of essentially the same processes present in the gel permeation chromatography of macromolecules. 

As has been discussed, size-exclusion chromatography (SEC) is a powerful tool for macromolecular separation and characterization. This technique separates macromolecules which vary in effective molecular size, utilizing columns that... 

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. 

Figure 12 Separation of Irganox 1076 and Irganox PS802 by size exclusion chromatography (duplicate solution injections of each antioxidant showing the reproducibility of the technique).

Size-exclusion chromatography, also termed gel-permeation or gel-filtration chromatography, separates proteins on the basis of their size and shape. As most proteins fractionated by this technique are considered to have approximately similar molecular shape, separation is often described as being on the basis of molecular mass, although such a description is somewhat simplistic. 

Size exclusion chromatography (SEC), also called gel permeation chromatography (GPC) or gel filtration chromatography (GFC), is a technique for separating dissolved species on the basis of their size. 

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