Synthesis of adsorbents with dendronic structures for protein hydrophobic interaction chromatography

Abstract

Here, we introduced a new technology based on the incorporation of dendrons—branched chemical structures—onto supports for synthesis of HIC adsorbents. In doing so we studied the synthesis and performance of these novel HIC dendron-based adsorbents. The adsorbents were synthesized in a facile two-step reaction. First, Sepharose 4FF (R) was chemically modified with polyester dendrons of different branching degrees i.e. third (G3) or fifth (G5) generations. Then, butyl-end valeric acid ligands were coupled to dendrons via ester bond formation. UV–vis spectrophotometry and FTIR analyses of the modified resins confirmed the presence of the dendrons and their ligands on them. Inclusion of dendrons allowed the increment of ligand density, 82.5±11 and 175.6±5.7μmol ligand/mL resin for RG3 and RG5, respectively. Static adsorption capacity of modified resins was found to be ∼60mg BSA/mL resin. Interestingly, dynamic binding capacity was higher at high flow rates, 62.5±0.8 and 58.0±0.5mg/mL for RG3 and RG5, respectively. RG3 was able to separate lipase, β-lactoglobulin and α-chymotrypsin selectively as well as fractionating of a whole proteome from yeast. This innovative technology will improve the existing HIC resin synthesis methods. It will also allow the reduction of the amount of adsorbent used in a chromatographic procedure and thus permit the use of smaller columns resulting in faster processes. Furthermore, this method could potentially be considered as a green technology since both, dendrons and ligands, are formed by ester bonds that are more biodegradable allowing the disposal of used resin waste in a more ecofriendly manner when compared to other exiting resins.