Towards a Universal LC-HRMS Method for Detecting Impurities in β-Lactam Antibiotics

M. Anzböck1, M. Kinzig2, F. Sörgel2, O. Scherf-Clavel1

1Ludwig-Maximilians-Universität München, Department of Pharmacy, Clinical Pharmacy and Pharmacotherapy, Butenandtstraße 5-13, 81377 München

2Institut für Biomedizinische und Pharmazeutische Forschung, Paul-Ehrlich-Straße 19, 90562 Nürnberg-Heroldsberg

 

Background

Piperacillin, in combination with the β-lactamase inhibitor tazobactam (Pip/Taz), is among the widely used β-lactam antibiotics for intravenous treatment of infections in hospitalized patients. During the COVID-19 pandemic, demand for this combination therapy increased significantly [1], contributing to repeated and documented supply shortages [2]. We aim to develop a generic workflow to screen beta-lactam antibiotics for known and unknown impurities as a tool for monitoring changes in active pharmaceutical ingredient (API) quality due to degradation during storage or changes in the supply chain.

Methods

We developed an untargeted workflow using liquid chromatography coupled to high-resolution mass spectrometry (qTOF). Known impurities were identified via certified reference standards according to the European Pharmacopoeia (Ph. Eur.) monograph [3]. As a proof-of-concept, a sample containing known related substances (Piperacillin for Peak Identification CRS) was used to set up the method. This sample was obtained from the European Directorate for the Quality of Medicines (EDQM). A stock solution was prepared in methanol at a concentration of 1 mg/mL and subsequently diluted to 100 µg/mL. Untargeted impurity profiling is performed using information-dependent acquisition (IDA). To identify signals originating from the analyte, the sample was analysed in parallel with a methanol blank under identical injection and matrix conditions.

Results
Multiple features were detected by the IDA workflow in both positive and negative ionization modes. In positive mode, three of the 18 known impurities listed in the Ph. Eur. were identified; in negative mode, one known impurity was detected. The identity of the others remains to be explored.

 

 

Conclusion

The IDA workflow appears suitable for the detection of unexpected impurities in beta-lactam antibiotics. However, further optimization is key to ensure a reliable detection of all known related substances and degradation products, thereby strengthening pharmaceutical quality control and contributing to long-term drug safety.

 

 

1.           Kern, W.V., et al., Impact of the COVID-19 Pandemic on Inpatient Antibiotic and Antifungal Drug Prescribing Volumes in Germany. Antibiotics (Basel), 2024. 13(9).

2.           BfArM, B.f.A.u.M. Veröffentlichte Lieferengpassmeldungen.  [cited 2025- 06-23]; Available from: https://anwendungen.pharmnet-bund.de/lieferengpassmeldungen/faces/public/meldungen.xhtml.

3.           EDQM, E.D.f.t.Q.o.M., Piperacillin sodium, in European Pharmacopoeia, E.P. Commission, Editor. 2023, Council of Europe: Strasbourg. p. 5549-5553.