Journal of Chromatography & Separation Techniques
Open Access
ISSN: 2157-7064
ISSN: 2157-7064
Vignesh Rajamanickam, Britta Eggenreich, Christoph Herwig and Oliver Spadiut
Research Division Biochemical Engineering, Institute of Chemical Engineering, Vienna University of Technology, Vienna, Austria
Posters & Accepted Abstracts: J Chromatogr Sep Tech
Escherichia coli is a well-studied recombinant host organism extensively used for recombinant protein production. Albeit E. coli is attributed with high product titres and growth on inexpensive media, overexpression of heterologous proteins often leads to accumulation of target protein in water-insoluble, misfolded aggregates called inclusion bodies (IBs). In practice, IBs are washed, solubilized and refolded to recover the target protein in its active form. Empirical complex washing and solubilization protocols are diverse and product specific and often render poor product quality with different impurities, which negatively impact subsequent refolding. Thus, fast and precise monitoring tools are needed to follow the impurity pattern along IB processing to judge the efficiency of each Unit Operation. However, such reliable monitoring tools are still scarce to date. In this study, we developed a novel toolbox using UV chromatogram fingerprints and chemometric techniques to monitor impurity pattern in IB washing and solubilization. Furthermore, we were not only able to monitor the process but also identify the optimal time point of transfer from one Unit Operation to the next. We are convinced that this toolbox will not only facilitate DSP monitoring, but will also allow enhanced process control in the future. The different unit operations where the novel toolbox was implemented is shown in Figure 1. Figure 1. Implementation of novel toolbox for monitoring different unit operations involved in inclusion body processing Keywords: process analytical technology, inclusion body processing, HPLC, process monitoring, Escherichia coli, chemometrics, chromatography
Vignesh Rajamanickam procured his Master of Science in pharmaceutical biotechnology from Hamburg University of applied sciences, Germany and, Bachelor of Technology in Biotechnology from Anna University, India. He started his PhD on March 2014 in Biochemical engineering from Vienna University of Technology (VUT), Austria. Currently, he is working as a project assistant for developing a novel PAT tool for bioprocess monitoring and control in Christian Doppler laboratory for mechanistic and physiological methods for improved bioprocesses, VUT, Austria.