Quantification of microplastic uptake and phytotoxicity in submerged aquatic plants using fluorescence spectroscopy

Microplastics (MPs) are persistent and ubiquitous contaminants in aquatic ecosystems, yet their interactions with submerged aquatic plants remain poorly understood. While MP-induced phytotoxicity has been extensively investigated in terrestrial plants, quantitative evidence for MP uptake and internal accumulation in submerged species is still limited. In this study, we investigated the phytotoxicity and accumulation patterns of fluorescent microplastics (FMPs) in two submerged aquatic plants, Bacopa lanigera and Rotala indica, using fluorescence spectroscopy. Plants were exposed to FMPs of two particle sizes (50 nm and 1 µm) across three exposure concentrations (0.001%, 0.01%, and 0.05%). Plant growth, chlorophyll content, fluorescence emission, and FMP accumulation were systematically evaluated. Our results demonstrated clear size- and concentration-dependent responses. Smaller particles (50 nm) showed significantly higher uptake and induced stronger phytotoxic effects than 1 µm particles, with pronounced growth inhibition and chlorophyll reduction observed at the highest concentration (0.05%). Fluorescence-based analysis enabled quantitative estimation of both surface-associated and internalized FMPs within plant tissues. Maximum surface accumulation reached 207 ppm, while internal (cross-sectional) accumulation reached up to 75 ppm, regardless of plant species. Under the respective experimental conditions, B. lanigera exhibited higher estimated FMP accumulation, whereas R. indica showed greater growth inhibition. These findings provide quantitative evidence of microplastic uptake and internal accumulation in submerged aquatic plants and highlight particle size as a critical determinant of phytotoxicity. Moreover, this study establishes a fluorescence-based methodological framework for estimating microplastic concentrations in aquatic plant tissues, contributing to improved ecological risk assessment of microplastics in freshwater ecosystems.