In planta localization of stilbenes within Picea abies phloem

Phenolic stilbene glucosides (astringin, isorhapontin, and piceid) and their aglycons commonly accumulate in the phloem of Norway spruce (Picea abies (L.) Karst.). However, current knowledge about the localization and accumulation of stilbenes within plant tissues and cells remains limited. Here, we used an innovative combination of novel micro-analytical techniques to evaluate stilbenes in frozen-hydrated condition, i.e., in planta, and freeze-dried condition across phloem tissues. Semi-quantitative TOF-SIMS (time-of-flight secondary-ion-mass-spectrometry) imaging in planta revealed that stilbenes were localized in axial parenchyma cells. Quantitative gas-chromatography analysis showed the highest stilbene content in the middle of collapsed phloem with decreases toward the outer phloem. The same trend was detected for soluble sugar and water content. The specimen water content may affect stilbene composition; the glucoside-to-aglycon ratio slightly decreased with decreases in water content. Phloem chemistry was correlated with 3D structures of phloem as analyzed by microtomography. The outer phloem was characterized by a high volume of empty parenchyma, reduced ray volume, and a large number of axial parenchyma with porous vacuolar contents. Increasing porosity from the inner to outer phloem was related to decreasing compactness of stilbenes, and possible secondary oxidation or polymerization. Our results indicate that aging-dependent changes in phloem may reduce cell functioning, which affects the capacity of the phloem to store water and sugar, and may reduce the defense potential of stilbenes in the axial parenchyma. Our results highlight the power of using a combination of techniques for evaluating tissue and cell-level mechanisms involved in plant secondary metabolite formation and metabolism.