Combining a tunable pinhole with synchronous fluorescence spectrometry for visualization and quantification of benzo[a]pyrene at the root epidermis microstructure of Kandelia obovata

The adsorption of polycyclic aromatic hydrocarbons (PAHs) by mangrove roots and their transport to chloroplasts is a potentially critical process that reduces the carbon sequestration efficiency of mangroves. Yet the crucial initial step, the distribution and retention of PAHs at the root epidermis microstructure, remains unclear. A novel method with a spatial resolution of 311 nm was developed for visualizing and quantifying benzo[a]pyrene (B[a]P) at the root epidermis microstructure (0.096 mm²) of Kandelia obovata (Ko). This method combined a tunable pinhole in laser confocal scanning microscopy with synchronous fluorescence spectrometry to reduce the auto-fluorescence interference in locating B[a]P and improve quantitative sensitivity. The linear range for the established method was 0.44–50.00 ng mm⁻², with a detection limit of 0.063 ng mm⁻² and a relative standard deviation of 9.45%. In a 60-day hydroponic experiment, B[a]P was primarily adsorbed along the epidermis cell walls of secondary lateral roots and lateral roots, with retained amounts of 0.65 ng mm⁻² and 0.49 ng mm⁻², respectively. It was found to cluster and predominantly accumulate at the epidermal cell surfaces of taproots (0.24 ng mm⁻²). B[a]P might enter inner root tissues through the root epidermal cell walls and surfaces of Ko, with the cell walls potentially being the main route. This study potentially provides a pathway for visualizing and quantifying B[a]P entering inner root tissues of mangroves.