GPC, using organic solvents, is used primarily for the analysis of polymers.

Separation occurs on the basis of decreasing molecular size, which to a first approximation equates to decreasing molecular weight. No chemical interaction must occur between the sample and the packing material.

A column is defined by its Exclusion Limit. Molecules bigger than this cannot enter the pores and pass unretained through the column. However columns with high exclusion limits offer very limited resolution of species with low molecular weight, so it is normal to require more than one column.

The total volume of the pores is known as the "Inner Volume" of the system, Vi. The largest peaks (at or above the Exclusion Limit) all elute in the Void volume of the system (V0). All peak will elute between V0 and V0+Vi. So it follows that the bigger the pore volume of the system, the better resolution can be achieved, and this is why GPC columns are usually wider and longer than normal HPLC Columns.

To separate a wide range of molecular sizes, there are two standard approaches:

1. Use a series of 3-4 different columns connected in series, each covering a different molecular weight range. Careful selection of the columns used (eg exclusion limits 400,000 + 70,000 + 5,000) will give a fairly linear calibration curve over the desired range.
2. Use 3 x Mixed bed columns. Effectively the packings from the three different column are mixed, and packed into the columns so that the samples pass through a continuous bed, and the separation occurs slowly through the columns. This gives a much better linearity and is always recommended where a suitable column is available.

Shodex have a range of columns to suit both of these approaches, and have also developed a third option (see next paragraph), unique to Shodex, which gives even better results. Mixed bed columns are designated with an M and a special version offering a really Linear calibration curve are designated with an L. Both the L and M versions are mixed bed columns, but the L version is a newer development, giveing a more linear calibration curve.

The latest approach has been achieved by developing a packing material with a wide pore-size distribution, so that it can cope with a wide molecular weight range using a single packing material. This gives a totally homogenous packing of the column, and the pore size distribution has been fine-tuned to give a truly linear calibration curve accross a wide range of molecular weights. We have a 17 page Technical Notebook from Shodex describing the use of this new column type, and these are available free of charge. To request a copy please send an email to gpcbooklet@laserchrom.co.uk, giving your name and company address, and we will be pleased to send you one by return.

Column format in GPC. Shodex offer three column formats for GPC:

1. The standard column size for GPC is 8.0 x 300mm. This is to achieve the large pore volume required, as described above. For most separations, this is the column size to choose.
2. Shodex also offer a Rapid Analysis format. These columns are 6.0mm id, and 150mm long. Analysis times are faster, because the column volmes are much lower. To combat the inherent reduction in resolution, these columns are offered with a smaller particle size packing.
3. Then there is the High Resolution Format. These columns are 4.6mm x 250mm. They use a very small particle size, yet are almost as long as a standard column, so with a slower flow rate the resolution is better than either of the other two. The smaller sample size means that a smaller flow cell can be used, thus developing almost 4 x the sensitivity of a standard column system.

Eluent choice for GPC is dependent upon several factors. Firstly, the sample must be soluble in the solvent used. This is not always as obvious as it seems, and often an elevated temperature is required to achieve total solubility. Secondly, the solvent ensures that there is no retention due to any interaction with the column other than that of size exclusion. And thirdly, since detection is usually by refractive index difference, the solvent should have as big a difference in RI from the samples as possible. Where possible, columns should be supplied in the solvent to be used, and hence Shodex GPC columns are offered in a range of solvents. Changing solvent with GPC columns is not as easy as with HPLC, and great care must be taken to follow the prescribed procedure.

Solvent Peak Separation Columns are available with some ranges of Shodex GPC columns. These are small pore size columns which cause the solvent used to dissolve the sample to elute a little after the end of the chromatogram. It also can improve the separation generally in the low molecular weight range. Further details can be found by following this link to the Shodex website: www.shodex.com

Guard Columns are used in GPC to prevent any chemical impurities in the polymer sample from binding chemically to the GPC column.