Ortho-, meta- and para-substituted fluorescent probes based on BODIPY for detecting esterase with application in...

Esterase plays a significant role in industrial production and life activities. The method of fluorescent probe is employed for detecting esterase due to their excellent performance. However, the restriction of recognition groups to specific positions severely limits the molecular design and performance optimization of fluorescent probe. Thus, three fluorescent probes (OBOD6, MBOD6 and PBOD6) based on BODIPY with recognition groups at ortho-, meta- and para- positions were designed and synthesized to systematically investigate the positional effects on performance of probes. All probes initially showed fluorescence quenching, followed by characteristic enhancement of fluorescent intensity at 587 nm (OBOD6), 577 nm (MBOD6) and 598 nm (PBOD6) upon esterase-catalyzed hydrolysis. Remarkably, OBOD6 exhibited the fastest reaction kinetics, nearly achieving conversion within 40 min at esterase of 0.6 U/mL. When incubated with esterase of 0.3 U/mL, MBOD6 (10 μM) nearly reached the reaction endpoint within 80 min, exhibiting the highest fluorescent enhancement and optimal detection sensitivity. MBOD6 could quantitatively detect esterase within a concentration range from 0.01 to 0.07 U/mL, with a detection limit of 2.23 × 10^-4 U/mL. Besides, machine learning was additionally applied to the quantitative detection of esterase using MBOD6. The model explained approximately 93.19% of the variation in esterase concentration, indicating good overall fitting capability. Finally, leveraging its outstanding performance, MBOD6 was successfully applied to imaging of esterase in both cells and zebrafish.