Improving sensitivity for the targeted LC-MS/MS analysis of the peptide bradykinin using a design of experiments approach.

Tanja Gangnus, Bjoern B Burckhardt

Talanta 2020 Oct 01

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The nonapeptide bradykinin is endogenously present only in low picomolar plasma concentrations, subsequently making reliable detection using liquid chromatography coupled to mass spectrometry (LC-MS/MS) challenging. Furthermore, non-specific adsorption during sample preparation and storage can lead to unpredictable peptide losses. To overcome these issues, a design of experiments (DoE) approach was applied, which consisted of a screening to identify impacting factors, optimisation and confirmation runs. On the one hand, different injection solvent compositions and sample collection materials were investigated in order to decrease non-specific adsorption. On the other hand, the addition of modifiers, which are known to enhance the signal intensity in LC-MS/MS, to the chromatographic mobile phase was examined. Polypropylene was the most suitable material among those investigated and resulted in a factor increase of 12.0 compared to LC-MS glass. The advantages of protein low-binding polypropylene versus standard polypropylene were fully compensated by the optimisation of the injection solvent. The latter substantially contributed to a decrease of non-specific adsorption of bradykinin. In this regard, bradykinin further benefitted from an organic fraction and a high amount of formic acid. Based on the DoE results, the final optimised injection solvent-consisting of 8.7% formic acid in 49.4/5.3/36.6 water/methanol/dimethyl sulfoxide (v/v/v)-was established. Furthermore, optimisation of the mobile phase composition yielded a signal intensity increase by a factor of 7.7. The transferability of the optimisation results conducted in neat solutions were successfully confirmed in human plasma. The applicability of this approach was further supported by the successful determination of low-abundance endogenous bradykinin levels in human plasma using LC-MS/MS. Copyright © 2020 Elsevier B.V. All rights reserved.

Citation

Tanja Gangnus, Bjoern B Burckhardt. Improving sensitivity for the targeted LC-MS/MS analysis of the peptide bradykinin using a design of experiments approach. Talanta. 2020 Oct 01;218:121134