Preparation of an internal surface reversed-phase restricted-access material for the analysis of hydrophobic molecules in biological matrices

•A novel method for preparation of reversed phase restricted access material (RAM).•The preparation method is simple and versatile.•The RAM has very good ability to retain small molecules and to exclude proteins.•RAM-SPE–HPLC method for detection of phthalate esters in milk was established.

Restricted-access materials (RAMs) have been broadly used for sample pretreatment in the chromatographic analysis of biological samples. In the present work, a novel internal surface reversed-phase (ISRP) RAM was prepared via surface-initiated atom transfer radical polymerization (SI-ATRP). Octadecyl and 4-(chloromethyl)phenyl were immobilized on silica using a one-pot synthesis approach to form a reversed-phase layer to retain small hydrophobic molecules, allowing the modified silica to serve as a macro-initiator. Then, poly(glycidyl methacrylate) (pGMA) was grafted onto the surface via SI-ATRP, and the epoxy groups were further hydrolyzed to form an external hydrophilic layer. The properties of this ISRP-RAM for the retention of small molecules and the exclusion of proteins were evaluated using solid-phase extraction (SPE). The removal efficiencies of bovine serum albumin (BSA) and lysozyme were 94.9% and 93.5%, respectively. The recoveries of five drugs, puerarin, p-hydroxybenzaldehyde, loratadine, nifedipine and diazepam, were 93.2–116%. Furthermore, the ISRP-RAM was employed for the SPE of five phthalate esters (PAEs) from bovine milk prior to high-performance liquid chromatography (HPLC) analysis. The results indicate that the prepared ISRP-RAM is qualified for practical bioanalysis.