Abstract: Molecularly imprinted solid-phase extraction (MISPE) microspheres were prepared using L-glutamic acid, 2-acrylamido-2-methyl propane sulfonic acid and N, N-methylene bis-acrylamide as template, functional monomer and cross-linker respectively and using the reversible addition-fragmentation chain transfer polymerization. Then the imprinted silica gel microspheres were characterized with Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM) and thermogravimetric analysis (TG). The results of the extraction experiment showed that MIPs exhibited excellent performance in separating glutamic acid from soy sauce, and an extraction capacity up to 140 μg/g was achieved. The rapid analysis of L-glutamic was realized by combining MISPE with high performance liquid chromatography technology. The linearity of L-glutamic acid was 1.47~58.9 μg/mL and the detection limit was 44.1 ng/mL. The recovery rate was 77.8%~82.2%.