Development and validation of liquid chromatographic method for the determination of lycopene in plasma Original Research Article

An isocratic reversed-phase liquid chromatographic method with spectrophotometric detection (472 nm) was developed and validated for the determination of lycopene in isolated lycopene crude material and in plasma. The method utilizes a Nova Pak C-18 column (3.9 mm×150 mm), 5 μm particle size, with a pre-column of the same type and a mobile phase consisting of methanol:acetonitrile:methylene chloride (55:30:15 (%, v/v)) at a flow rate of 1.0 ml min−1. The rentention time of lycopene was 4.8 min and the total run time was 8 min. Experiments were carried out at room temperature in the absence of direct sunlight. The isolation of lycopene from plasma was achieved with both liquid–liquid extraction (LLE) and solid-phase extraction (SPE) with recoveries of 94.5±5.1% and 62.3±2.6%, respectively. Calibration graphs from plasma standards were linear in the range 0.2–2.5 μM with a detection limit of 0.13 μM for the LLE procedure and 0.067–0.83 μM with a detection limit of 0.043 μM for the SPE procedure. Relative standard deviations were <5% for both procedures.