Raman spectroscopy resolves nitrogen-driven metabolic acclimation to urban air pollution in Quercus ilex

Urban air pollution may alter plant metabolism long before visible damage becomes apparent. Raman spectroscopy was evaluated as a rapid, non-destructive approach to resolve these biochemical adjustments. Mature Quercus ilex L. trees were sampled along a well-defined pollution gradient in Tuscany (Italy), spanning high, intermediate, and low levels of NO₂ and PM₁₀. Leaf Raman spectra revealed coordinated modulation of primary and secondary metabolism. Pigment-related bands (chlorophylls and carotenoids) increased toward the most polluted site, while inducible flavonoid signals showed site-dependent variation consistent with oxidative pressure in superficial tissues. These patterns were consistent with destructive biochemical analyses and chlorophyll fluorescence measurements, which indicated acclimation rather than photoinhibition damage. A composite Raman index showed a close site-level association with NO₂ exposure, suggesting that nitrogen-related urban pollution was the main exposure component linked to the observed metabolic response. Overall, Raman spectroscopy captures the chronic metabolic imprint of urban air pollution in Q. ilex, resolving coordinated pigment reinforcement and defensive activation without sample destruction. This approach provides a rapid and scalable framework for linking atmospheric chemistry to plant functional status in biomonitoring applications.