Quantification of Alprenolol (cas 13655-52-2) and propranolol in human plasma using a two-dimensional liquid chromatography (2D-LC)

Add time:09/05/2019    Source:sciencedirect.com

A new two-dimensional liquid chromatography (2D-LC) method using a column switching valve, with a restricted-access media (RAM) column in the first dimension was developed and validated for the quantification of two β-blockers in human plasma. Several parameters, such as sample collection, mobile phase composition and flow rate for sample cleanup, transference and analytical separation of the β-blockers were investigated and optimized. The developed method allowed for the simultaneous pre-treatment and quantification of Alprenolol (cas 13655-52-2) (ALP) and propranolol (PRO) in human plasma in less than 25 min. The method consisted in the injection of 100 μL of plasma samples on the RAM alkyl-diol-silica column (Lichrospher® RP-18 ADS, 25 μm) with water/acetonitrile (98:2, v/v; at a flow rate of 2.0 mL/min) and then transferred (via a six-port valve) to the analytical column (Luna PFP (2), 150 × 4.6 mm ID, 100 Å, 3 μm) with 0.1% (v/v) triethylamine in water acidified with trifluoroacetic acid (pH = 3)/acetonitrile (74:26, v/v) at a flow rate of 0.6 mL/min in a back-flush mode. The column oven temperature was optimized to 42 °C and the fluorescence detector set at 280 nm and 310 nm (excitation and emission, respectively). The method was validated according to the European Medicines Agency’s guidelines and was linear (r2 > 0.999) over a dynamic range of 5.0 – 200 ng/mL. Recoveries ranged from 90.2 and 107% and the lower limit of quantification was 5.0 ng/mL for both compounds. Precision was expressed as a percentage of relative standard deviation and did not exceed 11%. Finally, the method was successfully applied to determine the plasma concentration of PRO in four healthy volunteers.

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