The increased use of pesticides adversely affects the environment. To overcome this problem, there is a need to adopt sustainable agriculture. Cyperus rotundus L. is the main focus of the present work, which was carried out in order to find an effective and ecofriendly way to control this menacing weed. Allelopathic plants may show an enhancement of their phytotoxicity following their mixoploidisation. Hence, phytotoxic activity of diploid and mixoploid Trigonella foenum-graecum L. plants (fenugreek) harvested at vegetative stage was evaluated on C. rotundus. Treatments were applied either by pulverization of fenugreek aerial parts aqueous extracts (at 10, 20, and 30%) or by incorporation of the aerial parts powder into potting soil (in amounts of 0.2%, 0.6%, 1%), in pot experiments, under natural conditions. Results showed that phytotoxicity degree was dependent on the treatment and the ploidy level. Indeed, mixoploid fenugreek aqueous extract at 30% was the most harmful to weeds, whose leaves were totally burned, with 92.77% and 86% inhibition on root growth and weight, respectively. Pulverization by aqueous extracts of diploid plants had no significant effect on C. rotundus vegetative growth which was similar to control at all concentrations. The growth of C. rotun-dus, treated by adding mixoploid fenugreek powder to the potting soil, has been improved at the highest doses. However, its shoots and roots length and leaf number and weight were reduced by 45.45%, 63.88%, 66.66% and 74.54% respectively following adding diploid plants at 1%. As the pulverization treatment by mix-oploid plants was the most phytotoxic on C. rotundus, the reproducibility of this result was evaluated in field trial. Same to pot experiments, shoot growth was completely inhibited at 30%, with 91.51% and 92.17% reduc-tion in root growth and weight. Weed growth inhibition seems to be correlated with membrane deteriora-tion (proved by an increase in malondialdehyde (MDA) content by 67.55% and 57.85% in leaves and roots and electrolyte leakage increase by 64.88% and 46.46% in the same organs) and mitochondrial respiration disrup-tion due to a decrease in dehydrogenases activity. Chlorophyll and soluble sugars contents were similarly affected; while, proline and carotenoid contents increased.This study showed that mixoploidy could be an effective way to enhance the phytotoxicity of plants, which may be used to control the main weeds, often very resistant even to industrial herbicides. Understanding the different action mode of allelochemicals on target species may provide a basis for the development of bio-herbicides to boost up production in sustainable agriculture.(c) 2023 SAAB. Published by Elsevier B.V. All rights reserved.