Determination of estrogens and bisphenol A in bovine milk by automated on-line C30 solid-phase extraction coupled with high-performance liquid chromatography-mass spectrometry.

Using triacontyl bonded silica (C30) as on-line solid-phase extraction (SPE) material and a specially designed on-line analytical system which allowed large sample volume injection, a high speed and robust on-line SPE-HPLC-MS method was established for the analysis of five estrogens and bisphenol A (BPA) in milk samples. The milk sample is pretreated with acetonitrile for protein precipitation and then treated with primary secondary amine for the removal of polar impurities in the matrix. Then the pretreated sample can be automatically loaded by a LC pump. For effective extraction, an offshoot with NH4Ac solution of high-flow rate was employed to dilute the loaded sample by a mixing tee before sample was loaded onto the C30 extraction column. In this way, large volume injection (1 mL in this experiment) could be achieved. Some important parameters such as sample loading flow rate, sample dilution ratio and injection volume were optimized. Under the optimized conditions, the recoveries for all analytes range from 71.4 to 97.1% and reproducibility represented as RSDs are less than 15.0% (n=5) with milk samples spiked at 0.6 and 15 ng/mL of each analyte. To the authors' knowledge, it constitutes the first work describing a C30 on-line SPE-LC-MS analytical method for the screen and monitoring of these estrogens and BPA in milk.