Liquid chromatography?mass spectrometry approach for the characterization and purification of crude synthetic peptide hormones

Abstract

In this study, conditions for the optimal separation by LC-UV and characterisation by LC-ES-MS of crude mixtures generated during SPPS of several peptide hormones are compiled. The linear solvation energy relationship (LSER) methodology has been used to predict the retention of the target peptides and their side products and then to develop a separation LC methodology with applicability on both the analytical and preparative scale. Identification of these side products by LC-ES-MS analysis has been made on the basis of their calculated molecular masses. This method may be regarded as a key tool for the optimisation of the synthetic procedures and for complying with regulatory agencies' requirements before commercialisation of a safe and effective peptide-based pharmaceutical drug.

Improved Peptide Elution Time Prediction for Reversed-Phase Liquid Chromatography-MS by Incorporating Peptide Sequence Information

“…We have been working to apply additional information such as LC retention time to improve the confidence in peptide identifications, an approach also recently suggested by a group working to establish publication guidelines for peptide and protein identification . The use of peptide retention time information has proved useful in the past for LC method development of simple peptide mixtures, , purification of peptides of interest, , and identification of simple peptide mixtures in conjunction with UV and/or fluorescence and/or colorimetric methods of detection …”

Development and validation of a liquid chromatography–tandem mass spectrometry method for the determination of goserelin in rabbit plasma

“…Currently, several methods have been reported for the qualitative analysis of goserelin in a crude synthetic mixture using capillary zone electrophoresis (CZE)-MS [15] and LC-ESI-MS [11][12][13][14] and for its quantification in rat plasma using LC-Q-TOF [19]. However, due to long run times, low sensitivity, and insufficient method validation and sample preparation of these methods, a highly sensitive, selective, and rapid tandem mass spectrometry method (MS/MS) is necessary to quantify goserelin in biological fluids for pharmacokinetic studies.…”

“…Nevertheless, it has limitations due to the cross-reactivity with structurally similar peptides, which interfere with the precise quantitation of goserelin in plasma. Although a number of analytical methods for the qualitative or quantitative determination of goserelin, including radioimmunoassay [7,9,10], liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) [11][12][13][14], capillary zone electrophoresis (CZE)-UV/MS [15], multiple-injection CZE (MICZE) [16], CE hydrogen deuterium exchange-MS (CE-H/D-MS) [17], fast atom bombardment-MS (FAB-MS) [18], and quadrupole time-of-flight MS (Q-TOF MS) [19], have been published, these reports have usually been concerned with the crude synthetic peptide mixtures, degradation products, and pharmaceutical formulations. Michalet et al [19] used LC-Q-TOF tandem mass spectrometry for the quantitation of goserelin, and this is the only reported method, other than immunoassays, for goserelin analysis in biological fluids.…”

Expanding the Use of Dynamic Electrostatic Repulsion Reversed-Phase Chromatography: An Effective Elution Mode for Peptides Control and Analysis

“…Among all the techniques reported in the guidance, evaluating the related substance content in the manufactured peptide plays a central role. The most common method to achieve this target is reversed-phase chromatography (RPC), which exploits slight differences in hydrophobicity to separate analytes of interest [ 19 , 20 ].…”