Herein, a novel type of binary-metal-oxide-coated magnetic microspheres-titanium dioxide-zirconium dioxide-coated magnetic Fe 3 O 4 microspheres (Fe 3 O 4 @TiO 2 -ZrO 2 )-were prepared and employed to selectively capture phosphopeptides for the first time. The synthesized Fe 3 O 4 @TiO 2 -ZrO 2 magnetic functional microspheres were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy-dispersive X-ray (EDX) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), N 2 adsorption-desorption technique and magnetic measurements.Experimental conditions for trapping phosphopeptides were optimized by using b-casein as the standard protein. Under the optimal conditions, the mixture of b-casein and bovine serum albumin (BSA) with different proportions was considered as the semi-complex sample and non-fat milk as the real biological sample to evaluate the capability for trapping phosphopeptides. The results revealed that Fe 3 O 4 @TiO 2 -ZrO 2 not only concentrated more phosphopeptides than Fe 3 O 4 @TiO 2 or Fe 3 O 4 @ZrO 2 , but also enriched both the monophosphopeptides and multiphosphopeptides efficiently. The detection limit of this approach for the enrichment of phosphopeptides by Fe 3 O 4 @ZrO 2 -TiO 2 microspheres from the tryptic products of b-casein was 500 pM (500 mL).
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