Real-time fluorometric turn-on assay for protease activity and inhibitor screening with a benzoperylene probe.

A real-time fluorescence turn-on strategy for protease activity and inhibitor screening has been developed. A negatively charged benzo[ghi]perylene derivative (probe 1) was employed. Protamine is a cationic protein which can induce aggregation of probe 1 via strong electrostatic and hydrophobic interactions. The fluorescence of probe 1 was efficiently quenched. In the presence of a protease, protamine was enzymatically hydrolyzed and probe 1 de-aggregated. The recovery of the probe 1 monomer fluorescence could be detected. The protease activity could be monitored in real-time. In addition, upon addition of a protease inhibitor, the protease-catalyzed hydrolysis was inhibited, which led to a decreased fluorescence recovery. The fluorometric assay thus could also be employed for screening protease inhibitors.

Peroxidase activity of the coronene bisimide supramolecular architecture and its applications in colorimetric sensing of H2O2 and glucose.

A new water soluble coronene bisimide derivative (CTDI) was designed and synthesized. CTDI self-assembled in an aqueous solution and formed supramolecular nanofibers through π-π stacking and hydrophobic interactions. The nanofibers exhibit distinct peroxidase-like catalytic activity, and can catalyze the redox reaction of 3,3,5,5-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide. Clear assay solution color changes were observed. The peroxidase-like catalytic property was utilized for the sensitive detection of H2O2 and glucose. The assay shows excellent sensitivity, and 1 µM of glucose could be easily detected. Glucose detection in dilute human blood samples was also demonstrated, and the results were verified using a commercial glucose meter. Our method is simple, convenient, sensitive and selective, and could facilitate the sensing of glucose in relation to biological and biomedical research applications.

Polycation-induced benzoperylene probe excimer formation and the ratiometric detection of heparin and heparinase.

A benzoperylene probe excimer emission in an aqueous buffer solution is observed for the first time, and a novel ratiometric fluorescence method based on the probe excimer emission for the sensitive detection of heparin and heparinase is demonstrated. A negatively charged benzoperylene derivative, 6-(benzo[ghi]perylene-1,2-dicarboxylic imide-yl)hexanoic acid (BPDI), was employed. A polycation, poly(diallyldimethylammonium) chloride (poly-DDA), could induce aggregation of BPDI through noncovalent interactions. A decrease of BPDI monomer emission and a simultaneous increase of BPDI excimer emission were observed. Upon the addition of heparin, the strong binding between heparin and poly-DDA caused release of BPDI monomer molecules, and an excimer-monomer emission signal transition was detected. However, after the enzymatic hydrolysis of heparin by heparinase, heparin was hydrolyzed into small fragments, which weakened the competitive binding of heparin to poly-DDA. Poly-DDA induced aggregation of BPDI, and a monomer-excimer emission signal transition was detected. Our assay is simple, rapid, inexpensive, sensitive and selective, which could facilitate the heparin and heparinase related biochemical and biomedical research.