Phytoestrogen biomonitoring: an extractionless LC-MS/MS method for measuring urinary isoflavones and lignans by use of atmospheric pressure photoionization (APPI).

We present here a high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for quantifying phytoestrogenic isoflavones (daidzein, equol, genistein, and O-desmethylangolensin) and lignans (enterodiol and enterolactone) in urine without the use of extraction or the preconcentration techniques inherent in existing methods. The development of this concept was made possible by use of atmospheric pressure photoionization (APPI); an ionization technique that we found to improve analyte sensitivity relative to electrospray ionization and atmospheric pressure chemical ionization for this particular group of compounds. The analytical performance of this method was equal to or exceeded that of comparable methods. Between-run coefficients of variation (CVs) across three quality control (QC) pool levels analyzed in duplicate over 20 days were 3.1-5.8% CV; within-run CVs were 2.3-6.0%. Accuracy, as determined by average spike recovery in QC pools, was generally within ±10% of being quantitative (100%). Relative limits of detection were 0.04-0.4 ng/mL urine, with absolute detection limits as low as 0.1 pg. This method was applied to the analysis of >2,500 urine specimens for the 2005-2006 Centers for Disease Control and Prevention's National Health and Nutrition Examination Survey (NHANES). The method was capable of quantifying these compounds in 95-100% of study samples. This work is the first ever report of using APPI for the LC-MS/MS determination of these compounds in urine. It is also the first method of its kind to do so without any need for analyte extraction or preconcentration prior to analysis.