As industrial applications for polymers and biopolymers have expanded in recent years, the limitations of traditional characterisation techniques have become apparent. Size exclusion chromatography is widely used to determine the molecular weight distribution of a polymer, but su †ers from a number of drawbacks when used with ultra-high molecular weight (MW) polymers or polyelectrolytes. As a result, interest in the use of Ðeld Ñow fractionation (FFF) for the separation of macromolecules has grown.
FFF relies on a combination of Ðeld-driven and di †usive transport mechanisms to separate polymers in the range 103È1015 g mol~1 and/or particles of 5 nmÈ100 km, allowing MW and size distributions for virtually all macromolecules to be determined without the need for Ðltration. The open channel geometry minimises shear e †ects making it possible to separate fragile, high MW polymers, while the absence of a stationary phase means adsorption e †ects can be minimised. As a result, samples can be run in a mobile phase suitable for the polymer rather than selecting conditions to minimise column interactions which leads to the risk of aggregate formation in polyelectrolytes.
While it is possible to calculate the MW of a polymer eluting from an FFF channel using calibration techniques similar to those for size exclusion chromatography, the same problems are encountered as well. When analysing ultra-high MW polymers and/or gels, these are compounded by the need for calibration standards with a MW in excess of 106 g mol~1. The addition of multi-angle light scattering (MALS) to FFF has allowed absolute MW and size distributions to be obtained without the need for calibration, standards or assumptions. Complex mixtures of polymer, microgel, and macrogel can all be studied in a single run. By combining all of the information derived from FFFÈMALS, molecular conformation, density and branching levels can also be determined.
This paper outlines the principles of operation of FFFÈMALS, before moving on to discuss recent applications of the technique for the analysis of macromolecules and sub-micrometre particles.