Hydrophobic interaction chromatography for purification of monoPEGylated RNase A

Journal of Chromatography. a
Karla Mayolo-DeloisaJuan A Asenjo

Abstract

The chromatographic methods used for the purification of PEGylated proteins are mainly Size Exclusion (SEC) and Ion Exchange Chromatography (IEX). Although the PEGylation affects the protein hydrophobicity, Hydrophobic Interaction Chromatography (HIC) has not been extensively applied for the separation of these proteins. Purification of monoPEGylated Ribonuclease A (RNase A) using HIC is studied in this work. The products of the PEGylation reaction of RNase A with 20 kDa methoxy-poly(ethylene glycol) were separated using three resins with different degrees of hydrophobicity: Butyl, Octyl and Phenyl sepharose. The effects of resin type, concentration and salt type (ammonium sulphate or sodium chloride), and gradient length on the separation performance were evaluated. Yield and purity were calculated using the plate model. Under all conditions assayed the native protein was completely separated from PEGylated species. The best conditions for the purification of monoPEGylated RNase A were: Butyl sepharose, 1 M ammonium sulphate and 35 column volumes (CVs); this resulted in a yield as high as 85% with a purity of 97%. The purity of monoPEGylated RNase A is comparable to that obtained when the separation is performed using SEC, but...Continue Reading

References

Dec 5, 2006·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·M Elena LienqueoJuan A Asenjo
Sep 10, 2008·BioDrugs : Clinical Immunotherapeutics, Biopharmaceuticals and Gene Therapy·Francesco M Veronese, Anna Mero
May 8, 2010·Journal of Chromatography. a·Gopinath V AnnathurNatrajan Ramasubramanyan

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Citations

Jan 15, 2014·Journal of Chromatography. a·Jianlou NiuZhifeng Huang
Aug 19, 2014·Journal of Chromatography. a·Mayra Cisneros-RuizTodd M Przybycien
Nov 28, 2013·Journal of Molecular Recognition : JMR·Celeste C Ibarra-HerreraMarcelo Fernández-Lahore
Feb 6, 2015·Journal of Molecular Recognition : JMR·Karla Mayolo-DeloisaMarco Rito-Palomares
Oct 10, 2013·Journal of Separation Science·Xiaojiao ShangRaja Ghosh
Aug 27, 2014·Journal of Separation Science·Chunling GeTianwei Tan
Mar 29, 2016·Journal of Chromatography. a·Marco A Mata-GómezJosé González-Valdez
Feb 18, 2014·Journal of Controlled Release : Official Journal of the Controlled Release Society·David Pfister, Massimo Morbidelli
Feb 27, 2016·Biotechnology Progress·Karla Mayolo-DeloisaMarco Rito-Palomares
Aug 5, 2016·Journal of the American Chemical Society·Matthias FachPeter R Wich
Aug 16, 2016·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Jernej GašperšičNika Lendero Krajnc
Sep 18, 2020·Advances in Colloid and Interface Science·Shixiang LiuHailin Cong

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