Visual High-Throughput Screening for Developing a Fatty Acid Amide Hydrolase Natural Inhibitor Based on an Enzyme-Activated Fluorescent Probe.

Fatty acid amide hydrolase (FAAH) is an important drug target for the treatment of many disease related conditions such as pain, inflammation, and mood disorders due to its vital role in the metabolism of endocannabinoid. In our present work, a FAAH-activated fluorescent probe named THPO was developed, which possessed high selectivity and excellent sensitivity for FAAH in complex systems. Critically, its metabolite 7-amino-3H-phenoxazin-3-one (AHPO) has long excitation and emission wavelengths and high fluorescence quantum yield, which are necessary for monitoring the activity of FAAH in living systems. In addition, a visual high-throughput screening method for FAAH inhibitors was established using THPO, which resulted in the discovery of an efficient natural inhibitor Neobavaisoflavone that was identified from 68 traditional herbal medicines. These results indicated that THPO can be used as a molecular tool for the rapid evaluation of FAAH activity in complex systems as well as providing an effective approach to screen FAAH inhibitors and providing a boost for the discovery of therapeutic agents toward FAAH related diseases.