Laser-Induced Solid-Phase UV Fluorescence Spectroscopy for Rapid Detection of Polycyclic Aromatic Hydrocarbons in the Land Snail Bioindicator, Cantareus aspersus

Abstract

In ecotoxicological risk assessment, current methods for measuring the transfer and bioavailability of organic pollutants like polycyclic aromatic hydrocarbons (PAHs) in bioindicators are often destructive and environmentally unfriendly. These limitations are especially problematic when only small amounts of biological material are available. Here, we present a novel, high-throughput method combining laser-induced UV fluorescence spectroscopy (UV-LIF) and solid-phase spectroscopy (SPS) for rapid, in situ quantification of PAHs in land snails-a key bioindicator species. Using dual excitation wavelengths (266 nm and 355 nm), our method reliably detected pyrene and fluoranthene in snails exposed to varying concentrations, demonstrating clear dose-responses and inter-individual differences in bioaccumulation. The analysis time per sample was under four minutes. This approach allows simultaneous measurement of internal contaminant levels and health biomarkers in individual organisms and aligns with green chemistry principles. These findings establish a new, scalable tool for routine assessment of PAH transfer and bioavailability in diverse ecosystems.

Keywords: bioaccumulation; bioindicators; laser-induced UV spectroscopy; snails; solid-phase spectroscopy; transfers.

Figure 1

Experimental Workflow: Exposure, Sample Preparation, and Spectrometry Measurements (the control in step 1 refers to the sample without any PAH contamination). The negative control includes two sub-controls: one without solvents and one with solvents. The solvent control (ethanol) represents the sample in which the PAH solvent vector is added, but no PAHs are introduced, serving to assess the potential impact of the solvent. Each exposure modality was repeated in triplicate, with six snails placed in each replicate. For step 3, the different parts of the equipment are (1) beamsplitter, (2) mirror, (3) trigger (photodiode), (4) pierced mirror, (5) and (6) lens 1f, (7) high-pass filter, (8) slot, (9) shutter, (10) double motorized network (300 grt/mm–1200 grt/mm), (11) CCD 256 × 2048 px, (12) filter wheel, (13) signal transformer A > V, and (14) photomultiplier (PM) power supply.

Figure 2

Average fluorescence emission spectra of snail viscera for each exposure modality after excitation at 266 nm (A) and 355 nm (B). Fluorescence intensity is normalized in arbitrary units (au).

Figure 3

(A). Histogram of the 660 fluorescence maxima (110 per individual) at 395 nm after excitation at 355 nm, of the 6 individuals (a to f) of the PYR+FLT 200 condition. The vertical line represents the mean of the 6 median fluorescence maxima for each individual. (B). Fluorescence emission spectra of the 6 individuals in the ethanol condition and the 6 individuals in the PYR+FLT 200 condition after excitation at 355 nm. Fluorescence intensity is normalized in arbitrary units (au).

Figure 4

Comparison of the coefficient of variation (CV) for each condition in the pyrene peak at 395 nm, after excitation at 355 nm, as function of the red horizontal line representing the CV (%) for the control condition (Helinove without ethanol).

Figure 5

Medians of the fluorescence maxima for six individuals per experimental condition for the fluoranthene peak at 460 ± 10 nm (A,D), the pyrene peak at 370 ± 10 nm (B,E), and the pyrene peak at 395 ± 10 nm (C,F) after excitations at 266 nm and 355 nm, respectively. Boxplots represent the distribution of values: the central line corresponds to the median, the upper and lower bounds of the box indicate the first and third quartiles, whiskers extend to 1.5 times the interquartile range, and points outside this range are considered outliers. Statistical significance is indicated by asterisks (*), with * p < 0.05, ** p < 0.01, and *** p < 0.001. Fluorescence intensity is normalized in log10(arbitrary units (au) + 1) to facilitate comparisons between each modality and wavelength.