Spermine Sepharose as a clustered-charge anion exchange adsorbent

We previously showed that the affinity and capacity of ion exchange adsorbents of a given total charge density are improved by immobilization of the charges in pre-ordered clusters, rather than individually in random locations. This previous work used pentalysinamide and pentaargininamide as clustered ligands. This approach allows close control of cluster size, but is uneconomically expensive for some research and most practical applications. In this work, we demonstrate that the inexpensive synthetic analog of the natural polyamine spermine (H2NCH2CH2CH2NHCH2CH2CH2CH2NHCH2CH2CH2NH2) also can serve as the basis of effective clustered adsorbents. The calcium-depleted form of the protein α-lactalbumin, and RNA from bakerʹs yeast, were adsorbed on a spermine Sepharose adsorbent. This adsorbent exhibited enhanced α-lactalbumin binding capacity (Qmax > 1.6 and 1.3-fold higher than those for Qiagen DEAE and GE DEAE Sepharose adsorbents of much greater charge density) and higher initial binding affinity (Qmax/KD 2.4 and 2.1-fold higher, respectively). The new spermine-based matrix exhibited a higher value of the Z parameter, suggesting an increased number of apparent interaction sites between the protein and the resin, and functioned well in column mode.