SUMMARYLignin, one of the main structural polymer of plant cell walls, varies in amount and monomeric composition among tissue and cell types, as well as among plant species. However, few analytical methods are available that can conveniently and accurately determine the morphological distribution of lignin units at the cellular level. In this report, we used time-of-flight secondary ion mass spectrometry (TOF-SIMS) to directly map guaiacyl (G) and syringyl (S) lignin units in several successive growth rings of the maple xylem. TOF-SIMS imaging and a semiquantitative approach revealed clear difference in the annual distribution of lignins between the fiber and vessel. While the vessel walls were constantly G-rich with varied S/G ratios through a growth ring, the fibers showed fairly regular annual distribution of lignins in which the earlywood was S-rich with an almost constant S/G ratio and the latewood was G-rich resulting from a decrease of the S unit. The reliability of TOF-SIMS results was demonstrated by its high correlation with the results of thioacidolysis on radial distribution of the S/G ratio in several contiguous tree rings and also in the latewood and earlywood of each ring. These results indicate that TOF-SIMS allows direct visualization of lignin composition in plant tissues.

Field emission scanning electron microscopy was used to observe the inner surfaces of the developing secondary walls of earlywood tracheids of Abies sachalinensis Masters. Microfibrillar orientation in the secondary wall, as seen from the lumen side, changed in a clockwise direction from the outermost S1 to the middle of the S2 and from there counter-clockwise to the innermost S3. Sometimes microfibrils oriented in a steep S-helix were observed in the S3 layer. Lamellae showing different microfibrillar orientations in wall layers other than the S2 were observed beneath newly deposited microfibrils on the inner surface of the developing wall. Furthermore, on the inner surface of the wall forming the S12, S23 and S3, lamellae with microfibrils closely aligned at the same angle as one another and lacking spaces were not observed. These observations suggest that in layers other than the S2 most lamellae are not composed of closely spaced microfibrils.

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Abstract. The orientation of cellulose microfibrils (MFs) and the arrangement of cortical microtubules (MTs) in the developing tension-wood fibres of Japanese ash (Fraxinus mandshurica Rupr. var.japonica Maxim.) trees were investigated by electron and immunofluorescence microscopy. The MFs were deposited at an angle of about 45 ~ to the longitudinal axis of the fibre in an S-helical orientation at the initiation of secondary wall thickening. The MFs changed their orientation progressively, with clockwise rotation (viewed from the lumen side), from the S-helix until they were oriented approximately parallel to the fibre axis. This configuration can be considered as a semihelicoidal pattern. With arresting of rotation, a thick gelatinous (G-) layer was developed as a result of the repeated deposition of parallel MFs with a consistent texture. Two types of gelatinous fibre were identified on the basis of the orientation of MFs at the later stage of G-layer deposition. Microfibrils of type 1 were oriented parallel to the fibre axis; MFs of type 2 were laid down with counterclockwise rotation. The counterclockwise rotation of MFs was associated with a variation in the angle of MFs with respect to the fibre axis that ranged from 5 ~ to 25 ~ with a Z-helical orientation among the fibres. The MFs showed a high degree of parallelism at all stages of deposition during G-layer formation. No MFs with an S-helical orientation were observed in the G-layer. Based on these results, a model for the orientation and deposition of MFs in the secondary wall of tension-wood fibres with an $1 + G type of wall organization is proposed. The MT arrays changed progressively, with clockwise rotation (viewed from the lumen side), from an angle of about 35~40 ~ in a Z-helical orientation to an angle of approximately 0 ~ (parallel) to the fibre axis during G-layer formation. The parallelism between MTs and MFs was evident. The density of MTs in the developing tension-wood fibres Abbreviations: FE-SEM = field emission scanning electron microscopy; G = gelatinous layer; MF = cellulose microfibril; MT = cortical microtubule; $1 = outermost layer of the secondary wall; TEM = transmission electron microscopy Correspondence to: R. Funada; FAX: 81 (11) 736 1791 during formation of the G-layer was about 17-18 per gm of wall. It appears that MTs with a high density play a significant role in regulating the orientation of nascent MFs in the secondary walls of wood fibres. It also appears that the high degree of parallelism among MFs is closely related to the parallelism of MTs that are present at a high density.

Microspectrometry is the most definitive technique for obtaining both ultraviolet (UV) and visible light absorption spectra from a very limited area, and this technique allows the determination of lignin distribution throughout an individual cell wall. It is generally accepted that hardwood lignin .is composed mainly of guaiacyl and syringyl moieties. Our microspectrometric investigations revealed variation of lignin distribution within an annual ring in beech (Fagus crenata).