Hydrodynamic chromatography (HDC) in a packed column is a useful chromatogr
aphic mode for the rapid separation of polymers. This paper compared the re
tention of various circular double-stranded DNA molecules (3, 5 and 10 kbp)
and proteins (keyhole lumpet hemocyanin, ferritin, thyroglobulin, and cata
lase) in a chromatographic system using a C1 stationary phase and a acetoni
trile-phosphate buffer mixture as a mobile phase. For a similar molecular w
eight, the protein was retained more than the corresponding plasmid. This w
as attributed to the difference in the compactness between the two species.
As well, the retention dependence on the flow rate (0.03 to 1.5 mL/min) ex
hibited a different behavior in relation to the type of biopolymer.
The protein retention curve showed a decrease in the relative retention tim
e until 0.2 mL/min, followed by an increase for the high size polymers, onl
y, while the plasmid retention increased over the entire flow rate range. T
his observation confirmed that the migration in HDC was dependent on two an
tagonist phenomena, i.e. stress induced diffusion and polymer deformation.
This work demonstrated that the HDC separation of protein was optimal for a
flow rate equal to 0.2 mL/min.