Bioanalytical method development and validation for the determination of metoprolol and meldonium in human plasma

Abstract

Aim.The main purpose of this study was to develop a simple, precise, rapid and accurate method for the quantification of metoprolol and meldonium in human plasma. Materials and methods. The resolution of peaks of metoprolol was best achieved with Discovery C18, 50 × 2.1 mm, 5 μm column and meldonium - ZORBAX HILIC Plus, 50 × 2.1 mm, 3.5 μm column. Samples of metoprolol were chromatographed in a gradient mode (eluent A (acetonitrile – water – formic acid, 5 : 95 : 0.1 v/v), eluent B (acetonitrile – formic acid, 100 : 0.1 v/v)). The initial content of the eluent B is 0%, which increases linearly by 1.0 min to 100% and to 1.11 min returns to the initial 0%. The mobile phase was delivered at a flow rate of 0.400 mL/min into the mass spectrometer ESI chamber. The injection volume was 5μl. Samples of meldonium were chromatographed in a isocratic using mobile phase water – acetonitrile – ammonium formate buffer 200 мМ, 20 : 75 : 5 v/v). Results.The total chromatographic run time was 2.0 minutes and the elution of metoprolol, meldonium and IS occurred at ~1.39 and 1.18 minutes, respectively.A linear response function was established at 2 - 200 ng/mL for metoprolol and 50 -5000 ng/mL for meldonium in human plasma. The% mean recovery for metoprolol in LQC, MQC and HQC was 99.0%, 107.5% and 96.8%, for meldonium in LQC, MQC and HQC was 94.1%, 100.2% and 93.1% respectively. The lowest concentration with the RSD <20% was taken as LLOQ and was found to be 2.31 ng/mL for metoprolol, 47.70 ng/mL for meldonium. The % accuracy of LLOQ samples prepared with the different biological matrix lots were found 115.4% for metoprolol and 95.5% for meldonium, which were found within the range of 80.00–120.00% for the seven different plasma lots. % CV for LLOQ samples was observed as 12.8% and 7.7% respectively, which are within 20.00% of the acceptance criteria. Conclusion.A rapid method was developed for simultaneous determination of metoprolol and meldonium in human plasma. The method was strictly validated according to the ICH guidelines. Acquired results demonstrate that proposed strategy can be effortlessly and advantageously applied for routine examination of metoprolol and meldonium in human plasma.