Directly visualizing the spatial distribution of exogenous humectants in a leaf matrix via stimulated Raman scattering imaging.

Plant leaves are not only vital for material exchange with the environment, but also act as primary sites for the deposition, permeation, and action of exogenous functional molecules in agricultural applications such as pest control, nutrient supplementation, and post-harvest processing. Visualizing the spatial distribution of exogenous molecules in plant leaves is of great importance. Traditional methods like chromatography and mass spectrometry may require destroying samples and make it difficult to obtain in situ distribution information. Typical Raman spectroscopy suffers from fluorescence interference and low sensitivity. This study uses tobacco leaves as a model and applies stimulated Raman Scattering (SRS) microscopy combined with Lasso spectral unmixing to achieve label-free, high-resolution, rapid visualization, and semi-quantitative analysis of propylene glycol and glycerol. The results show that SRS can clearly distinguish the characteristic spectra of tobacco, propylene glycol, and glycerol, visually presenting the distribution differences of polyols on the surface and inside the leaf. Besides, the co-localization of the two components and the analysis of their relatively mass ratios are also realized. This method provides a universal technical solution for detecting the distribution of exogenous small molecules in plant leaves.