Microscopic spatiotemporal changes in cell wall cellulose and pectin during Nicotiana tabacum L. leaf growth and senescence based on label-free Raman microspectroscopic imaging combined with multivariate curve resolution

The plant cell wall, composed mainly of polysaccharides, lignin, and structural proteins, supports the architecture, mechanics, and functions of plants. Developing appropriate chemical imaging methods to study spatiotemporal changes of cell wall structural components at the microscopic level is important for understanding plant growth and senescence. In this study, tobacco (Nicotiana tabacum L.), a widely cultivated economic crop and model plant, was selected as the research object. Based on Raman confocal imaging combined with a multivariate curve resolution model, a label-free, in situ, high-throughput and high specificity imaging method for cellulose, high methylated pectin, and low methylated pectin in tobacco leaf cell wall was established to study their microscopic spatiotemporal changes during leaf growth and senescence (flue-curing) processes. The results based on the proposed method revealed that cellulose and pectin levels in the midrib cell wall gradually increased as the leaves matured, from appeared mainly at the cell corners and middle lamella respectively, to appeared in the cell corners, middle lamella, and cell wall. The same trend was observed in the lateral vein cell walls, where cellulose and pectin levels gradually increased. During the flue-curing process, cellulose and highly methylated pectin degraded. The proposed chemical imaging method is expected to provide a label-free, in situ, and high-throughput cell imaging technique for investigating the microscopic spatiotemporal distribution of the main structural components of the leaf cell wall.