Inhibitory mechanism of O-methylated quercetins, highly potent β-secretase inhibitors isolated from Caragana balchaschensis (Kom.) Pojark.

ETHNOPHARMACOLOGICAL RELEVANCE

Caragana has a standing history of implementation in Traditional Chinese Medicine (TCM). Most species of this genus have been explored for multi-functional purposes, such as promoting blood circulation and curing neuralgia, fatigue, migraine, arthritis, and vascular hypertension (Meng et al., 2009). Among them, the well-known species C. sinica showed the most promising potential to increase the expression of ADAM10 among 313 tested medicinal plants, which is one of the promising approach for the treatment of Alzheimer's disease (AD). (Schuck et al., 2015).

AIM OF THIS STUDY

The aim of this work is to explore β-secretase inhibitory activity of compounds isolated from the aerial part of endemic Caragana balchaschensis (Kom.) Pojark. We provided a full characterization of their inhibitory mechanisms, binding affinities, and binding modes.

MATERIALS AND METHODS

The isolation of quercetin derivatives was accomplished by various chromatographical approaches and their structures were annotated by spectroscopic analysis. The detailed kinetic behavior of β-secretase inhibitors was determined by estimation of kinetic parameters (K, V, K, and K). Binding affinities (K) and binding modes of inhibitors were elucidated by fluorescence quenching and molecular docking studies, respectively.

RESULTS

O-methylated quercetins (2-7) were significantly effective in β-secretase inhibition with IC ranging from 1.2 to 6.5 μM. The most active one (6) was 20-fold effective than the mother skeleton, quercetin. The O-methyl motif was a critical factor in β-secretase inhibition: tri-O-methylated (1.2 μM) > di-O-methylated (3.5 μM) > mono-O-methylated (6.5 μM) > quercetin (25.2 μM). In the kinetic study, all quercetins (1-7) showed a noncompetitive inhibition, but glucoside ones (8 and 9) were mixed type I inhibitors. The binding affinities (K) were agreed with inhibitory potencies. The O-methylated quercetins were annotated as the most natural abundant metabolites in the aerial part by LC-ESI-TOF/MS. Binding modes of inhibitors to enzyme were elucidated by molecular docking experiments.

CONCLUSION

This study disclosed that most of the major phenolic metabolites of the aerial part of C. balchaschensis are O-methylated quercetins, which have a significant inhibitory effect on β-secretase, which is a critical factor for AD.