A novel fluorometric assay for trypsin on the basis of a gemini anionic surfactant/BSA/NR supramolecular assembly system with favorable salt resistance

Abstract

A novel label-free fluorometric assay for trypsin detection was successfully established on the basis of an anionic sulfonate gemini surfactant C₁₂C₃C₁₂(SO₃)₂/bovine serum albumin (BSA)/nile red (NR) supramolecular assembly system through encapsulation of NR as the fluorescence probe in the hydrophobic interior of the C₁₂C₃C₁₂(SO₃)₂/BSA complex. The optimized conditions for the formation of this supramolecular assembly system were composed of 5 μM C₁₂C₃C₁₂(SO₃)₂, 0.3 mg mL⁻¹ BSA and 1 μM NR. The hydrolysis of BSA into short peptides was facilitated by trypsin addition, which resulted in the dissociation of the assembly system and fluorescence quenching. The quenching efficiency linearly increased over the trypsin concentration of 1.6–150 ng mL⁻¹ with a limit of detection of 1.6 ng mL⁻¹. The supramolecular assembly system showed favorable salt resistance and remained stable in NaCl solution below 200 mM. Moreover, this supramolecular assembly system can also be employed for the detection of trypsin in human urine with a recovery efficiency of 95.2%–103.8%.