Synergistic effects of titanium dioxide nanoparticles and microplastics on lentil seeds by a non-invasive biospeckle optical coherence tomography

Titanium dioxide nanoparticles (TiO₂ NPs) are used in agriculture, cosmetics, energy, and environmental applications, necessitating advanced methods to evaluate their effects on biological systems such as plant growth. This study demonstrates the use of biospeckle optical coherence tomography (bOCT), a novel and non-invasive technique, to rapidly assess the size and concentration-dependent impacts of TiO₂ NPs as well as the synergistic effects of TiO₂ NPs and polyethylene microplastics (PEMPs) on lentil (<i>Lens culinaris)</i> seeds. The primary objective was to validate bOCT as a rapid and non-invasive tool for assessing NPs-induced biological responses in plants. Seeds were treated with TiO₂ particles (<5 µm,<100 nm, 21 nm) at concentrations of 0, 25, 100, and 200 mg/L. The sizes were selected based on commercially available TiO₂ NPs grades in industrial and agricultural applications. For synergy experiments, TiO₂ NPs (21 nm) at 25 and 100 mg/L were combined with PEMPs (744-4990 nm) at concentration-based ratios of 1:1 and 1:2. A swept-source OCT system (central wavelength: 1.3 µm; bandwidth: 125 nm; sweep frequency: 20 kHz), acquired OCT structural images at 12.5 frames per second and biospeckle images were calculated as the ratio of the standard deviation to the mean of 100 OCT structural images over an 8-second interval, at 0, 5, 10, and 20 hours (h) post-exposure. Smaller TiO₂ NPs (<100 nm) enhanced internal activity at lower concentrations (25 mg/L), while larger particles (<5 µm) exhibited similar effects at higher concentrations (200 mg/L). These observations were qualitatively consistent with conventional physiological measurements, including germination rates, growth parameters, and antioxidative enzyme activities recorded over 7 days. The co-application of TiO₂ NPs and PEMPs at a 1:1 ratio alleviated the reduction in internal activity caused by PEMPs alone, while the 1:2 ratio led to a significant decrease in biospeckle contrast, indicating suppressed internal seed activity. bOCT successfully detected early biological responses of TiO₂ NPs within 20 h, demonstrating its efficiency compared to the conventional methods. The ability of bOCT to monitor dynamic internal changes highlights its potential as a rapid tool for assessing NPs as well as their synergistic effects with polyethylene microplastics on plants.