Size exclusion materials

Ghijs, M., DeWaele, C., and Sandra, R, Experiments with size exclusion material in microchromatography. Part 1. Microcolumn SEC, ]. HRC, 13, 651, 1990. 

Proteins and glycoproteins Sephadex G-15 G-200, Bio-Gel P-6 -P-lOO, other polymer-based size-exclusion material and their DEAE-bonded forms, CIS on large-pore (300A) support and CIS on polymer-based material (e.g., HEMA and Hamilton PRP)... 

Two classes of micron-sized stationary phases have been encountered in this section silica particles and cross-linked polymer resin beads. Both materials are porous, with pore sizes ranging from approximately 50 to 4000 A for silica particles and from 50 to 1,000,000 A for divinylbenzene cross-linked polystyrene resins. In size-exclusion chromatography, also called molecular-exclusion or gel-permeation chromatography, separation is based on the solute s ability to enter into the pores of the column packing. Smaller solutes spend proportionally more time within the pores and, consequently, take longer to elute from the column. 

Validation Considerations. Mechanisms other then size exclusion maybe operative ia the removal of vimses from biological fluids. Thus vims removal must be vaUdated within the parameters set forth for the production process and usiag membrane material representative of the product line of the filter. 

Conductivity detectors, commonly employed in ion chromatography, can be used to determine ionic materials at levels of parts per million (ppm) or parts per bUHon (ppb) in aqueous mobile phases. The infrared (ir) detector is one that may be used in either nonselective or selective detection. Its most common use has been as a detector in size-exclusion chromatography, although it is not limited to sec. The detector is limited to use in systems in which the mobile phase is transparent to the ir wavelength being monitored. It is possible to obtain complete spectra, much as in some gc-ir experiments, if the flow is not very high or can be stopped momentarily. 

Rcnsl dipcptidflsc (from porcine kidney cortex) [9031-96-3] Mr 47,000 [EC 3.4.13.11]. Purified by homogenising the tissue, extracting with Triton X-100, elimination of insoluble material, and ion-exchange, size exclusion and affinity chromatography. [Hitchcock et al. Anal Biochem 163 219 7957.]... 

Microspherical polymer beads are widely used as packing materials for chromatography and a variety of other applications. Size exclusion chromatography is based on pore size and pore-size distribution of microbeads to separate... 

Size exclusion was first noted in the late fifties when separations of proteins on columns packed with swollen maize starch were observed (Lindqvist and Storgards, 1955 Lathe and Ruthven, 1956). The run time was typically 48 hr. With the advent of a commercial material for size separation of molecules, a gel of cross-linked dextran, researchers were given a purposely made material for size exclusion, or gel filtration, of solutes as described in the classical work by Porath and Flodin (1959). The material, named Sephadex, was made available commercially by Pharmacia in 1959. This promoted a rapid development of the technique and it was soon applied to the separation of proteins and aqueous polymers. The work by Porath and Flodin promoted Moore (1964) to apply the technique to size separation, gel permeation chromatography of organic molecules on gels of lightly cross-linked polystyrene (i.e., Styragel). 

Modern SEC columns are packed with material other than polystyrene gels, such as porous silica particles or highly cross-linked styrene-divinylbenzene copolymers. Because of improvements in speed and resolution, the term SEC is sometimes replaced by the term high-performance size-exclusion chromatography (HPSEC). 

Most size exclusion chromatography (SEC) practitioners select their columns primarily to cover the molar mass area of interest and to ensure compatibility with the mobile phase(s) applied. A further parameter to judge is the column efficiency expressed, e.g., by the theoretical plate count or related values, which are measured by appropriate low molar mass probes. It follows the apparent linearity of the calibration dependence and the attainable selectivity of separation the latter parameter is in turn connected with the width of the molar mass range covered by the column and depends on both the pore size distribution and the pore volume of the packing material. Other important column parameters are the column production repeatability, availability, and price. Unfortunately, the interactive properties of SEC columns are often overlooked. 

The elution of such gels is an example not of size exclusion but rather of hydrodynamic fractionation (HDF). However, it must be remembered that merely being able to physically fit an insoluble material through the column interstices is not the only criterion for whether the GPC/HDF analysis of an insoluble material will be successful. A well-designed HDF packing and eluant combination will often elute up to the estimated radius in Eq. (5), but adsorption can drastically limit this upper analysis radius. For example, work in our laboratory using an 8-mm-bead-diameter Polymer Laboratories aqueous GPC column for HDF found that that column could not elute 204 nM pSty particles, even though Eq. (5) estimates a critical radius of —1.5 jam. 

A. Packing Materials for Size Exclusion Chromatography (SEC) and HOPC... 

Chemically attached copolymers of iV-vinylpyrrolidone (N-VP) and N-(2-hydroxyethyl)acrylamide (N-HEAA) steeply decrease the inherent glass adsorp-tivity which is observed for proteins in aqueous buffer solutions. Thus, it became possible to apply the prepared materials to the size exclusion chromatography of viruses and ribosomes. 

---