Pyrrolizidine alkaloids (PAs) are one of the largest distributed classes of toxins in nature. They have a wide range of toxicity, such as hepatotoxicity, pulmonary toxicity, neuronal toxicity, and carcinogenesis. Yet, biological targets responsible for these effects are not well addressed. Using methods of computational biology for target identification, we tested more than 200 PAs. We used a machine-learning approach that applies structural similarity for target identification, ChemMapper, and SwissTargetPrediction. The predicted targets with high probabilities were muscarinic acetylcholine receptor M1. The predicted interactions between these two targets and PAs were further studied by molecular docking-based binding energies using AutoDock and VinaLC, which revealed good binding affinities. The PAs are bound to the same binding pocket as pirenzepine, a known M1 antagonist. These results were confirmed by in vitro assays showing that PAs increased the levels of intracellular calcium. We conclude that PAs are potential acetylcholine receptor M1 antagonists. This elucidates for the first time the serious neuro-oncological toxicities exerted by PA consumption.

This study investigated the effects of zinc oxide nanoparticles (ZnO-NPs) and salicylic acid (SA) on artemisinin production in Artemisia annua L., the compound with bioactivity against various diseases, including malaria. A range of methods have been suggested to increase artemisinin production. This research focused on enhancing artemisinin content through elicitor application. Six- week-old plants were subjected to foliar spray at different dose of nanoparticles and SA over a 96 h period. Results demonstrated that ZnO-NPs act as a stronger elicitor compared to SA; specifically, expose to 200 mg L^–1 ZnO-NPs for 96 h significantly increased the artemisinin content by 43.3% relative to the control. Gene expression analysis revealed significant upregulation of AaWRKY1, AaMYB2, HMGR, and CYP71AV1 in response to ZnO-NPs, mediated likely through H₂O₂ signalling. Concurrently, ZnO-NPs enhanced total protein content and induced antioxidant and defence-related enzymes including SOD, CAT, POX, and PAL. In contrast, 1 mM SA modestly upregulated AaWRKY1, AaMYB2, and HMGR, but had negligible impact on CYP71AV1. This is the first report demonstrating that ZnO-NPs, as effective elicitor, do not induce oxidative damage, but instead stimulated a protective antioxidant response that facilitate redirection of metabolic flux toward enhancing the yield of pharmaceutically important secondary metabolite such as artemisinin.