Downstream Processing of Therapeutic Peptides by Means of Preparative Liquid Chromatography

Abstract

The market of biomolecules with therapeutic scopes, including peptides, is continuously expanding. The interest towards this class of pharmaceuticals is stimulated by the broad range of bioactivities that peptides can trigger in the human body. The main production methods to obtain peptides are enzymatic hydrolysis, microbial fermentation, recombinant approach and, especially, chemical synthesis. None of these methods, however, produce exclusively the target product. Other species represent impurities that, for safety and pharmaceutical quality reasons, must be removed. The remarkable production volumes of peptide mixtures have generated a strong interest towards the purification procedures, particularly due to their relevant impact on the manufacturing costs. The purification method of choice is mainly preparative liquid chromatography, because of its flexibility, which allows one to choose case-by-case the experimental conditions that most suitably fit that particular purification problem. Different modes of chromatography that can cover almost every separation case are reviewed in this article. Additionally, an outlook to a very recent continuous chromatographic process (namely Multicolumn Countercurrent Solvent Gradient Purification, MCSGP) and future perspectives regarding purification strategies will be considered at the end of this review.

Sustainability in peptide chemistry: current synthesis and purification technologies and future challenges

“…22). 215,229 To alleviate this trade-off, it is possible to decrease the sample volume loaded or to use less steep gradients, but this would come at the cost of lower productivity and higher solvent consumption. 230 This is why the downstream processing represents the manufacturing bottleneck for many biopharmaceuticals.…”

An Integrated Chromatographic Strategy for the Large-Scale Extraction of Ergosterol from Tulasnellaceae sp.

“…Preparative high-performance liquid chromatography (prep-HPLC) is an efficient strategy for the large-scale separation of individual constituents from complex samples, including microorganisms and natural products [15][16][17][18]. This method is extensively employed in numerous areas because of its excellent column efficiency, superior separation reproducibility, online detection, and automatic control [19,20].…”

Preparation of mixed-mode stationary phase for separation of peptides and proteins in high performance liquid chromatography

“…However, RP stationary phases do not provide satisfactory separation of peptides and proteins due to their complex structure and amphoteric nature 14 , 15 . Therefore, specially designed stationary phases are required for the analysis of peptides and proteins that have both polar and nonpolar moieties to interact with these analytes and retain them 16 . Mixed-mode chromatography which offers multimodal interaction could be an alternative to RP-LC for the separation of peptides, proteins and other complex mixtures.…”