Fluorescence Spectroscopic Studies to Evaluate Binding Interaction between Hoechst 33258 and Bilirubin

Area of Science:

• Biophysical Chemistry
• Molecular Interactions
• Fluorescence Spectroscopy

Background:

• Bilirubin (Br) is a key marker for jaundice, and understanding its interactions with biological molecules is crucial.
• Hoechst33258 (H258) is a fluorescent dye with potential applications in molecular detection.

Purpose of the Study:

• To investigate the binding mechanism between the fluorophore Hoechst33258 (H258) and bilirubin (Br) using spectroscopic methods.
• To elucidate the nature of the quenching interaction and its implications for using H258 as a probe.

Main Methods:

• Detailed spectroscopic studies including fluorescence, absorption, and lifetime measurements.
• Förster Resonance Energy Transfer (FRET) analysis and molecular docking simulations.
• Application of the Stern-Volmer model and sphere of action model for quenching analysis.

Main Results:

• Concentration-dependent fluorescence quenching of H258 by Br was observed.
• Analysis indicated a combination of static and dynamic quenching mechanisms.
• FRET and lifetime studies confirmed dynamic quenching, with an efficiency of 59% and a reduced H258 lifetime.
• Molecular docking revealed non-covalent interactions, including hydrogen bonding between H258 and Br, supported by HSA.

Conclusions:

• Hoechst33258 (H258) exhibits both static and dynamic quenching interactions with bilirubin (Br).
• The binding involves hydrogen bonding and Van-der-Waals forces, consistent with spectroscopic and computational findings.
• H258 shows potential as an efficient fluorophore for monitoring binding interactions and detecting bilirubin.