Ecotoxicological bioassays with terrestrial plants: a holistic view of standards, guidelines, and protocols.

Terrestrial and aquatic ecosystems face various chemicals that might induce acute and/or long-term harm. To assess these impacts, ecotoxicological bioassays are essential. However, bioassays using animals, particularly mammals, are costly, time-consuming, and raise ethical concerns. In this context, terrestrial plants emerge as a viable alternative to conventional assays. Thus, the aim of this review was to address the history and evolution of plant bioassays, highlighting the main regulations, guidelines, and protocols governing the use of terrestrial plants in ecotoxicological tests. Initially, plant bioassays were employed to assess the cytogenotoxic effects of chemicals, gaining prominence with the GENE-TOX program in the 80s. Subsequently, plants were used in allelopathy bioassays and in studies aimed to examine the ecotoxicity of pesticides in soil. Currently, ecotoxicological bioassays with plants are regulated by specific standards, such as ASTM E1963-22, EPA 600/3-88/029, EPS 1/RM/45, ISO 11269-1, ISO 11269-2, ISO 17126, ISO 18763, ISO 29200, ISO 22030, OECD-208, OECD-227, OCSPP 850.4100, OCSPP 850.4230, OCSPP 850.4800 and OPPTS 850.4200. The existing protocols standardize bioassays in greenhouse and lab environments, and the duration of the tests varies from hours to months. The main ecotoxicological parameters to be analyzed after exposure include germination percentage, survival rate, root length, aerial part length, fresh mass of exposed plants, and phytotoxicity symptoms. In addition, the absorption rate of substances and genotoxic and mutagenic effects might also be assessed. Therefore, data in this review demonstrate that terrestrial plants represent an important tool in the analysis of environmental risks associated with chemicals and might serve as crucial allies in modern ecotoxicology.