Silicas SynChropak

SynChropak GPC supports were introduced in 1978 as the first commercial columns for high-performance liquid chromatography of proteins. SynChropak GPC columns were based on research developed by Fred Regnier and coworkers in 1976 (1,2). The first columns were only available in 10-yu,m particles with a 100-A pore diameter, but as silica technology advanced, the range of available pore diameters increased and 5-yu,m particle diameters became available. SynChropak GPC and CATSEC occasionally were prepared on larger particles on a custom basis, but generally these products have been intended for analytical applications. 

SynChropak size exclusion supports are composed of spherical uniformly porous silica that has been derivatized with a suitable layer. SynChropak GPC supports are available in six pore diameters ranging from 50 to 4000 A and particle diameters from 5 to 10 /zm. SynChropak CATSEC supports are available in four pore diameters. Table 10.1 details the physical characteristics of the product lines. 

SynChropak GPC supports are bonded with y-glycidoxypropylsilane by a proprietary process that results in a thin, neutral hydrophilic layer that totally covers the silanol sites of the silica. The silica backbone prevents the supports from swelling. 

The other restriction of SynChropak size exclusion columns is a general one for silica-based supports, that of pH. The most harmful pH is that above 7.5 due to silica dissolution. The bonded phase of SynChropak GPC has some polymeric properties therefore, it is not removed rapidly from the silica at pH 2-3. The bonded phase of SynChropak CATSEC is polymeric and stable at pH 2-7.5. 

Scientific (Northbrook, IL) contain a silica support with a -y-glycidoxypropylsi-lane-bonded phase to minimize interaction with anionic and neutral polymers. The columns come in five different pore sizes ranging from 100 to 4000 A. The packing material has a diameter from 5 to 10 /cm and yields in excess of 10,000 plate counts. With a rigid silica packing material, the columns can withstand high pressure (maximum of 3000 psi) and can be used under a variety of salt and/or buffered conditions. A mobile phase above pH 8, however, will dissolve the silica support of the column (21). A summary of the experimental conditions used for Synchropak columns is described in Table 20.8. 

Mercury poroslmetry data of these packings are given In Table IV. It Is of Interest to note that the pore-size distribution of CPG Is significantly more narrow than that of Syn-Chropak, a surface-modified porous silica (LlChrospher). These different physical characteristics may help to explain the existence of micropores In SynChropak. Because of the wide pore-size distribution of this packing. It seems reasonable that this material also contains a population of micropores which are only accessible to D2O. In mercury poroslmetry measurements, the lower pore size limit Is about 30A. 

From these studies with SynChropak SEC packings and controlled porosity glass, it is concluded that the silica packing contains a population of micropores which are differentially accessible to low molecular weight probes of total permeation volume. It is not known, however, if the microporosity in the 100 and 300A SynChropak SEC packings is the result of the rather wide pore-size distribution and whether all silicas contain micropores. 

Figure 9. Size-exclusion calibration for silica sols column, two 25 X 0.49 cm Synchropak-1000 mobile phase, 5 mM triethanolamine, pH 7.5 flow rate, 0.50 mL/min sample, 0.020 raL, 0.6%in mobile phase and UVdetector, 240 nm.

Porous silica Zorbax PSM Li Chrospher Li Chrospher Diol Synchropak GPC Chromegapore Sphere sil p-Bondagel E Protein Column TSK Type SW... 

Manufacturers or suppliers of silica-based packings for SEC are Beckman (UltraSi ierogel), ffio-Raid (Bio-Sil SEC), Mac-Mod (Zorbax PSM and GF), Merck (Lkhrospher), Syncbrom (SynChropak GPQ, Showa Denko (Protein KW), Tosoh (TSKgel SW), and Waters (Protein-Pak). 

Pfannkoch et compared the electrostatic characteristics of a number of derivatized silica packings via the elution of citrate ion in pH 7 phosphate buffer. One may view the relative decrease in Ksec foi" citrate as the ionic strength is decreased from 2.4 M to 0.026 M as a measure of the effect of SiO" groups. The order of citrate exclusion measured in this way was Waters 1-125 (0.44) < TSK SW 2000 (0.49) < SynChropak 6PC 100 (0.51) < LiChrosob Diol... 

SynChropak was the first commercial bonded silica of this type, followed by LIChrosorb DIOL and LiChrospher DIOL. Protein-Pak materials as well as Sl-Polyol are similar in surface composition. Recently a specifically pH-stabIIized, glycerol-bonded silica has been introduced on the market Zorbax Bio-Series 6F 250. The most popular are the TSK gel SW types which are thought to be bonded with a glycerol-ether moiety. On annealing from 400 to 823 K, the TSK gel SW 3000 showed a weight loss of ca. 15X while the carbon content was 7.3X, Indicating on oxygen-rich... 

The majority of reports in the literature concerning SEC of peptides and proteins tend to describe chromatographic conditions designed to ensure a pure size exclusion process, although it is often overlooked that the nonideal properties of size exclusion columns can be advantageous in the separation of peptides [37,77]. Excellent examples of this were reported by Yasukawa et al. [78], who separated various hydrophilic and hydrophobic dipeptides on a non-silica-based column by isocratic elution with 50 mM sodium phosphate buffer (pH 7.0). The peptides, retained by adsorption, were eluted in the order of increasing hydrophobicity. Similarly, Mant et al. [37] demonstrated that the separation of five peptide standards of 10, 20, 30, 40, and 50 residues was markedly improved on a SynChropak GPC-60 column when advantage was taken of the nonspecific adsorptive properties of the column rather than when the column was utilized under conditions of ideal SEC. Thus, what is occasionally viewed as a column limitation may become a useful analytical tool. 

SynChropak SEC supports (SynChrom Inc., Lafayette, USA), and also Aquapore OH columns (Brownlee Labs, USA), are composed of silica with a glycerylpropyl t/)... 

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