Design and Synthesis of Novel Indole-Derived N‑Methylcarbamoylguanidinyl Chitinase Inhibitors with Significantly Improved Insecticidal Activity

Chitinases play an important role in the molting process of insects and are potential targets for the development of green insecticides. Based on the feature that the +1/+2 sites in OfChtI, OfChtII, and OfChi-h have tryptophan residues in mismatch-parallel position, a strategy to introduce indole scaffold into chitinase inhibitors was proposed, and multitarget chitinase inhibitors containing N-methylcarbamoylguanidinyl and indole scaffold were successfully synthesized. The inhibitory activity showed that compound 8u exhibited significant inhibitory activity against OfChtI, OfChtII, and OfChi-h, with IC50 values of 0.7, 0.79, and 0.58 μM, and K i values of 0.05 ± 0.005, 0.065 ± 0.004, and 0.025 ± 0.006 μM, respectively. In vivo insecticidal activity showed that compounds 8a and 8g exhibited excellent insecticidal activity against Plutella xylostella and Mythimna separata, with LC50 values of 0.79 and 9.17 mg/L against P. xylostella, respectively, and 3.58 and 83.09 mg/L against M. separata, respectively, making them the most potent chitinase inhibitors with in vivo insecticidal activity discovered to date. The inhibition mechanism and binding free energy results suggested that N-methylcarbamoylguanidinyl binds to the −1 catalytic site, while additional interactions acquired by π–π stacking and hydrophobic interactions of the indole scaffold with tryptophan increase the binding affinity of the targets to chitinases. This work provides a new direction for the development of chitinase inhibitors with compounds 8a and 8g potentially serving as promising candidates for pesticide development.