Molecularly imprinted silica nanoparticles (SP‐MIP) are synthesized for the real‐time optical detection of low‐molecular‐weight compounds. Azo‐initiator‐modified silica beads are functionalized through reversible addition‐fragmentation chain transfer (RAFT) polymerization, which leads to efficient control of the grafted layer. The copolymerization of methacrylic acid (MAA) and ethylene glycol dimethacrylate (EDMA) on azo initiator‐coated silica particles (≈100 nm) using chain transfer agent (2‐phenylprop‐2‐yl‐dithiobenzoate) is carried out in the presence of a target analyte molecule (l‐Boc‐phenylalanine anilide, l‐BFA). The chemical and morphological properties of SP‐MIP are characterized by scanning electron microscopy, X‐ray photoelectron spectroscopy, Brunauer–Emmett–Teller surface analysis, and thermogravimetric analysis. Finally, SP‐MIP is located on the gold surface to be used as a biorecognition layer on the surface plasmon resonance spectrometer (SPR). The sensitivity, response time, and selectivity of SP‐MIP are investigated by three similar analogous molecules (l‐Boc‐Tryptophan, l‐Boc‐Tyrosine, and l‐Boc‐Phenylalanine) and the imprinted particle surface showed excellent relative selectivity toward l‐Boc‐Phenylalanine (l‐BFA) (k = 61), while the sensitivity is recorded as limit of detection = 1.72 × 10−4 m.