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Accurate quantification of impurities in pure peptide material – angiotensin I: Comparison of calibration requirements and method performance characteristics of liquid chromatography coupled to hybrid tandem mass spectrometry and linear ion trap high‐resolution mass spectrometry
Author(s) -
Stoppacher N.,
Josephs R. D.,
Daireaux A.,
Choteau T.,
Westwood S.,
Wielgosz R. I.
Publication year - 2015
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.7261
Subject(s) - chemistry , chromatography , mass spectrometry , impurity , electrospray ionization , liquid chromatography–mass spectrometry , tandem mass spectrometry , angiotensin ii , protein mass spectrometry , analytical chemistry (journal) , peptide , capillary electrophoresis–mass spectrometry , selected reaction monitoring , biochemistry , organic chemistry , receptor
Rationale The prohormone angiotensin I (ANG I) [amino acid sequence: DRVYIHPFHL] and other structurally related peptide hormones play an essential role in the regulation of the water and electrolyte balance in the human body as well as blood pressure. ANG I is a biomarker for hypertension and diabetes. Therefore, well‐characterized pure reference materials and comparable and SI‐traceable analytical characterization methods are required to establish reference measurement systems (RMS) for laboratory medicine. Methods Two analytical characterization methods based on liquid chromatography/mass spectrometry (LC/MS) systems with electrospray ionization have been developed and validated in‐house. Both high‐resolution MS (hrMS) and hybrid‐tandem MS/MS were used for the identification and quantification of the major structurally related peptide impurities of ANG I. The impurities were quantified by use of external calibrations with original impurity standards. Mass fraction impurity values and corresponding expanded measurement uncertainties were calculated. Results Five structurally related degradation products were detected as major impurities in a 'pure' ANG I material. The peptides ANG (2‐10) [RVYIHPFHL], ANG II [DRVYIHPF] and three ANG I isomers [DRVY L HPFHL, DRVYIHPFH I and DRVY L HPFH I ] were identified and corresponding mass fraction values calculated that range from 0.66 to 4.86 mg/g. Conclusions The mass fraction values for the major related peptide impurities in the ANG I material obtained with both LC/hrMS and LC/MS/MS systems are in excellent agreement. This study emphasizes the importance of mass spectrometric techniques for application to mass balance approaches for mass fraction value and uncertainty assignment of impurities in 'pure' substance reference materials for peptides. Copyright © 2015 John Wiley & Sons, Ltd.