Effects of iron oxide nanoparticles on phenotype and metabolite changes in hemp clones (Cannabis sativa L.)

● Fe₃O₄ NPs increased the biomass and chlorophyll content of hemp clones. ● Fe₃O₄ NPs penetrated and were internalized by root cells. ● Fe₃O₄ NPs induced the alteration of metabolite profiles in hemp leaves. ● The psychoactive compound THC in hemp leaves was significantly down-regulated. We investigated the effect of iron oxide nanoparticles (Fe₃O₄ NPs, ~17 nm in size) on the phenotype and metabolite changes in hemp (Cannabis sativa L.), an annual crop distributed worldwide. Hemp clones were grown in hydroponic cultures with Fe₃O₄ NPs (50, 100, 200, or 500 mg/L) for four weeks. TEM and ICP-MS were used to determine Fe₃O₄ NPs uptake and translocation. LC-MS-based metabolomics was employed to explore the deep insight into the effect of Fe₃O₄ NPs on hemp plants. The results revealed that plant growth enhanced gradually with increasing concentrations of given NPs up to 200 mg/L, which improved the fresh weight and dry weight by 36.13% and 74.68%, respectively, compared to the control. Even at a high dose (500 mg/L), Fe₃O₄ NPs promoted plant growth, including increased biomass and tissue length. NPs significantly increased the iron and chlorophyll content in plant tissues Increased catalase activity and reduced hydrogen peroxide content in hemp leaves suggested that the Fe₃O₄ NPs activated the defense system. TEM showed that NPs were abundantly attached to the cell wall and dispersed throughout the root cells. Metabolomics revealed that Fe₃O₄ NPs induced metabolic reprogramming in hemp leaves, including the up-regulation of carbohydrates and organic acids, and down-regulation of antioxidants, especially tetrahydrocannabinol (THC). The significantly up-regulated metabolites, including peonidin and 2-hydroxycinnamic acid, could be involved in photosynthesis in hemp plants. These results demonstrate the potential of Fe₃O₄ NPs for promoting hemp growth and decreasing the THC content at low doses.