Two different hydrophilic materials with large pores, Superose 6 and F
ractogel EMD BioSec (S), which are designed for size-exclusion chromat
ography (SEC) of plasma proteins with high molecular masses, are teste
d for their performance on a preparative scale. The model mixtures are
preparations of the clotting factors VIII (FVII) and IX (FIX). A comb
ination of a Fractogel EMD BioSec (S) column and a Superose 6 column h
as proved to be particularly effective for separations in a wide molec
ular size range, from several millions down to about 20 000. Superose
6 showed good results on a small scale as well as on a large scale, ev
en in the molecular mass range over 1 000 000. However, recovery of FV
III clotting activity was less than 70% with this material and therefo
re not satisfactory. Fractogel did not perform well in terms of separa
tion on a small scale, However, in the case of biopolymers with high m
olecular masses, separation was improved by using larger columns. With
Fractogel, recovery of activity of the two clotting factors FVIII and
FIX was satisfactory, above 80%. On a large scale, the active fractio
n in the clotting factor concentrate was successfully separated from t
he non-active fraction with either size exclusion (SE) material. In th
e preparation under investigation, the clotting factor VIII is found i
n a complex with the von Willebrand factor (vWF). The FVIII-VWF comple
x has a molecular mass of several millions. It dissociates in the pres
ence of high concentrations of Ca2+ ions. Under such conditions FVIII
and VWF were successfully separated with both SEC columns. (C) 1998 El
sevier Science B.V.