Detection of polycyclic aromatic hydrocarbons (PAHs) in Medicago sativa L. by fluorescence microscopy

Alfalfa (Medicago sativa L.) leaves and roots grown in the presence and absence of 150ppm polycyclic aromatic hydrocarbons (PAHs) for 40days are washed with distilled water, dried with filter paper and immersed in n-hexane for 30s to remove adhered surface PAHs. Subsequently, the samples are mounted on slides with 50% glycerol and viewed under a fluorescence microscope (Leica DM 5000B model with Leica Filter cube A: UV excitation range, excitation filter BP340–380nm, dichromatic mirror 400nm and suppression filter LP 425nm). Images are recorded with a digital camera (Leica DFC 500) under UV and visible light. The location of PAHs is evidenced by the detection of blue autofluorescence, typical of the PAH studied, under UV light (*). This is the first report of PAHs in alfalfa tissues detected by fluorescence microscopy and intense fluorescence in the glandular secreting trichomes (GSTs) of plants grown in contaminated soil. These trichomes, with as as-yet-unknown function, may be sites of PAH conjugation and degradation in alfalfa. [Display omitted] •First study to detect the presence of PAHs in aerial and root tissues of Medicago sativa L. using fluorescence microscopy.•First study to propose a physiological function of PAH conjugation and/or degradation trichomes (GSTs) of M. sativa L.•Simple, rapid and effective method for detecting PAHs, organic pollutants, derived from oil and oil products in M. sativa L. Green technologies, such as phytoremediation, are effective for removing organic pollutants derived from oil and oil products, including polycyclic aromatic hydrocarbons (PAHs). Given the increasing popularity of these sustainable remediation techniques, methods based on fluorescence microscopy and multiphoton microscopy for the environmental monitoring of such pollutants have emerged in recent decades as effective tools for phytoremediation studies aimed at understanding the fate of these contaminants in plants. However, little is known about the cellular and molecular mechanisms involved in PAH uptake, responses and degradation by plants. Thus, the present study aimed to detect the location of pyrene, anthracene and phenanthrene using fluorescence microscopy techniques in shoots and roots of Medicago sativa L. (alfalfa) plants grown in artificially contaminated soil (150ppm PAHs) for 40days. Leaflet and root samples were then collected and observed under a fluorescence microscope to detect the presence of PAHs in various tissues. One important findi