Development and Validation of a Simple High Performance Liquid Chromatography/UV Method for Simultaneous Determination of Urinary Phthalates in Human Urine

This study aimed to develop a simple HPLC method for the simultaneous determination of four urinary phthalate (PAE) metabolites, including monomethyl PAE (MMP), monoethyl PAE (MEP), mono-n-butyl PAE (MBP) and mono (2-ethylhexyl) PAE (MEHP). The metabolites were simultaneously analyzed by a simple dilution method with acetonitrile and phosphate buffer pH 2.0 followed by filtration. No interfering peaks were observed at the retention time of the investigated analytes. Separation of four PAE metabolites was successfully achieved in less than 16.5 min with good resolution. The linear range of calibration curves for the four analytes were respectively from 10-3,000 ng/mL for MMP and MEP, 20-3,000 ng/mL for MBP and MEHP with correlation coefficient >= 0.998. Intra- and inter-day precision ranged from 0.5 to 10.2% and 2.1 to 9.9%; whereas intra- and inter-day relative errors (%bias) were less than 11.1 and 7.3%, respectively. This method was successfully applied to preliminary screen four PAEs in urine samples from Thai children and adolescents. Since, the levels of PAEs metabolites have been found to associate with health conditions including precocious puberty in children. Measurement of their metabolites can be helpful in clinical application and can provide the basis for appropriate risk management measures.