Development and validation of an assay for the measurement of gentamicin concentrations in dried blood spots using UHPLC-MS/MS

Abstract

Gentamicin sulfate (GEN) is an aminoglycoside antibiotic with a narrow therapeutic range of plasma concentrations. The collection of venous blood represents a significant burden for patients, especially in neonatology. Dried blood spots (DBS) obtained from capillary blood can be an alternative for drug measurements in this particular population. This study aimed to develop and validate an assay for the quantification of GEN in DBS using UHPLC-MS/MS. Total GEN concentrations were obtained by adding the individual concentrations of the GEN forms C1, C1a, and C2. The assay used a DBS disk containing approximately 17 μL of blood for GEN quantitation in the range of 0.1–40 mg L-1. Measurement accuracy for total GEN was in the range of 102.6–108.6%, inter-assay precision was 11.3–13.1% and intra-assay precision was 9.1–12.8.% GEN was stable for 21 days at − 20 and 8 °C, but only for 24 h at room temperature. Blood Hct affected the accuracy within acceptable limits (93.8–95% at Hct% of 30, 104.3–113% at Hct% of 50). Blood spotted volume did not affect GEN measurement accuracy. Concentrations of GEN in DBS obtained after heel pricks were correlated to plasma levels in a small cohort of neonatal patients. However, percentual differences between estimated plasma concentrations and actual plasma levels presented values between − 64–35.3% (average difference of − 1.9%). The use of DBS for the measurement of GEN concentrations can increase access to TDM of this antibiotic due to the ease of sample collection and the facilitated specimen transportation logistics when testing is not available onsite.