An SEC/MALS study of alternan degradation during size-exclusion chromatographic analysis.
Ultrahigh-molar-mass (M) polymers such as DNA, cellulose, and polyolefins are routinely analyzed using size-exclusion chromatography (SEC) to obtain molar mass averages, distributions, and architectural information. It has long been contended that high-M polymers can degrade during SEC analysis; if true, the inaccurate molar mass information obtained can adversely affect decisions regarding processing and end-use properties of the macromolecules. However, most evidence to the effect of degradation has been circumstantial and open to alternative interpretation. For example, the shift in SEC elution volume as a function of increased chromatographic flow rate, observed using only a concentration-sensitive detector, may be the result of degradation or of elution via a nondegradatory slalom chromatography mechanism. Here, using both concentration-sensitive and multiangle static light-scattering detection, we provide unambiguous evidence that the polysaccharide alternan actually degrades during SEC analysis. The decrease in molar mass and size of alternan with increasing flow rate, measured using light scattering, allows ruling out an SC mode of elution and can only be interpreted as due to degradation. These findings demonstrate the extreme fragility of ultrahigh-M polymers and the care that must be taken for accurate characterization.
Striegel
André M
AM
Department of Chemistry and Biochemistry, Florida State University, Tallahassee, FL 32306-4390, USA. striegel@chem.fsu.edu
Isenberg
Samantha L
SL
Côté
Gregory L
GL
D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002850:Chromatography, Gel / Q000379:methods*; D005936:Glucans / Q000032:analysis*; D008027:Light; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010316:Particle Size; D012542:Scattering, Radiation; D012680:Sensitivity and Specificity
D005936:Glucans; C439358:alternan