Duropore pvdf centrifugal filters

Ten microliters of 2000 nM class-specific stable isotope surrogates and ~1000nM unlabeled pesticide analytes were spiked into hexane-rinsed 50 mL borosilicate screw-threaded conical glass tubes, to which 1 mL of pooled breastmilk (n=4) or water negative control (n=2) were added. Contents were vortexed for approximately 2 seconds. Twenty milliliters of 2:1 (v/v) hexane:dichloromethane were added to all tubes. The tubes were capped with Teflon-lined caps, sonicated for 15 minutes and vortexed for 3 minutes. The samples were centrifuged at 3500 rcf (g) at 4°C for 15 minutes to separate the phases. The top hexane:dichloromethane layer was transferred to a second tube and the extraction repeated. The supernatant of the second extraction was pooled with the first one. Total supernatants were brought to dryness by nitrogen evaporation. The residues were reconstituted in 100μL of 200nM CUDA/PHAU internal standard solution, vortexed for 3 minutes at room temperature and chilled in wet ice for 15 minutes. The extracts were transferred to 0.1um Millipore Duropore PVDF centrifugal filters (cat # UFC30VV00), centrifuged for 2 minutes at 4500g and 4°C, transferred to a 150μL glass insert in a 2 mL amber autosampler vial with a slit cap (Waters Corp, Milford, MA), and analyzed by UPLC-MS/MS.

Ten microliters of 2000nM class specific stabile isotope surrogates and ~1000 nM analyte spike mixture were spiked into hexane-rinsed 13 × 100 mm glass tubes with polypropylene screw-top caps. One-hundred microliters of pooled breastmilk (n=4) or water (n=2) were added and contents were vortexed for 2 seconds. Two milliliters of a 2:1 hexane:dichloromethane solution were added to all tubes, which were then capped and sonicated for 15 minutes then, vortexed for 3 minutes. Samples were centrifuged at 3500 rcf (g) at 4°C for 15 minutes to separate the phases. The top layer was transferred with a glass Pasteur pipette to a second clean tube and the extraction was repeated, adding supernatant to the second vial. Total supernatants were brought to dryness by centrifugal vacuum. Residues were reconstituted in 100μL of 200nM CUDA/PHAU internal standard solution, capped and vortexed for 3 minutes at room temperature, and chilled in wet ice for 15 minutes. Extracts were transferred to 0.1um Millipore Duropore PVDF centrifugal filters, centrifuged for 2 minutes at 4500g and 4°C, then transferred to a 150μL glass insert in a 2 mL amber autosampler vial with a slit cap (Waters Corp, Milford, MA), and analyzed by UPLC-MS/MS.