Sensitive determination of gentamicin in plasma using ion-exchange solid-phase extraction followed by UHPLC-MS/MS analysis

Abstract

Background: Therapeutic drug monitoring (TDM) of gentamicin sulfate (GEN) is usually recommended, particularly in critical patients. Only a few reports had described the determination of GEN in plasma or plasma using LC-MS/MS. Objective: This study aimed to develop and validate a sensitive ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) assay for the quantification of GEN in small volumes of human plasma. Results: The use of a very low concentration of the ion-pairing agent HFBA allowed significant retention of the very polar GEN forms in a reversed phase UHPLC column. The solid-phase extraction (SPE) procedure allowed clean extracts, with no interferences detected in blank samples, and high sensitivity. The assay was linear on the range of 0.2–40 mg L−1 of GEN complex. The combined GEN complex had inter-assay CV of 8.8–10.0%, intra-assay CV of 10.2–11.0%, and accuracy of 96.8–104.0%. The assay was applied to 17 clinical samples obtained from neonate patients. Measured concentrations were in the range of 0.15–3.57 mg L−1 for GEN C1, 0.12–3.55 mg L−1 for GEN C1a, 0.20–5.77 mg L−1 for GEN C2, and 0.47–12.88 mg L−1 for the GEN complex, all within the linear range of the assay. Conclusion: A sensitive assay for the quantification of gentamicin in plasma using anion-exchange SPE and UHPLC-MS/MS was validated. The assay can be used for TDM of gentamicin, particularly in centers with access to proper instrumentation and with a low demand for gentamicin measurements, where immunoassays are not cost-effective.

Keywords

• Gentamicin
• UHPLC-MS/MS
• Solid-phase extraction
• Therapeutic drug monitoring
• Neonates