Hydration effects on spacing of primary-wall cellulose microfibrils: a small angle X-ray scattering study

Celery collenchyma cell walls are typical of primary plant cell walls in their composition but contain unusually well-oriented cellulose microfibrils that are packed with more regularity than normal, permitting small-angle X-ray scattering (SAXS) experiments that would not otherwise be possible. Small-angle scattering data were obtained for the cell walls in essentially their native state and for isolated cellulose, in a fibrous form that retained the physical shape and microfibril orientation of the native cell walls. The scattering patterns showed a distinct peak attributed to the interference contribution to the convolution of form and interference functions. The position of the peak attributed to the interference function implied a mean centre-to-centre microfibril spacing of approximately 3.2 nm in dry isolated cellulose and 3.8 nm in dry cell walls. Hydration increased the mean microfibril spacing in the cell walls to 5.4 nm but had only a small effect on the mean microfibril spacing of isolated cellulose. In the scattering profile from intact, hydrated cell walls it was just possible to discern the position of the first Bessel minimum, from which a microfibril diameter in the range 3.1–3.6 nm may be estimated. This estimate is likely to include attached hemicellulose chains. Porod plots of scattering intensity indicated a relatively sharp interface between microfibrils and their immediate surroundings. The SAXS data imply that cellulose microfibrils 2.6–3.0 nm in diameter are not quite in lateral contact with one another in the isolated cellulose and are augmented by hemicelluloses and separated by readily hydrated matrix polysaccharides in the native plant cell wall.     

Elucidation of the fibrous structure of <Emphasis Type="Italic">Musaceae</Emphasis> maturate rachis

In the last years several composites and high performance materials with woody and non-woody natural fibers have been developed. In this study, a morphological study of agricultural residues as rachis from Musaceae plants cultivated in Colombia has been carried out. Fibrous structures as fiber bundles, elementary or ultimate fibers and even cellulose microfibrils grouped together into microfibril bundles have been observed. Both biological retting and chemical procedures like alkali treatments combined with alkaline peroxide and acid addition have been used. Different microscopic techniques as optical (OM), confocal (CM), scanning electron (SEM), and atomic force (AFM) ones have been used for analysis of isolated samples. A hierarchical arrangement from conducting tissues and fiber bundles to cellulose microfibrils in Musaceae rachis has been noted. All of these structures can be isolated by biological and chemical processes at the corresponding arrangement level. This means that Musaceae rachises constitute a source of new interesting biodegradable raw materials with multiple possibilities in dimensions and morphologies for several industries. A strong presence of crystal structures exists on fiber surfaces, being their occurrence related to the maturate state of rachis samples. Additionally, a top-down scheme is proposed for understanding the structuration of rachis at each length scale.     

Comparison of the characteristics of cellulose microfibril aggregates of wood, rice straw and potato tuber

The focus of this study has been to isolate cellulose microfibril aggregates by the one-time grinding treatment from wood, rice straw and potato tuber, and to compare their morphological and mechanical properties. Field emission scanning electron microscopy images showed that the diameter range of isolated microfibril aggregates from wood, 12–20 nm, was smaller than those from rice straw and potato tuber, 12–35 nm and 12–55 nm, respectively. These differences were observed even in the purified rice straws and potato tuber before the grinder treatment, but were hardly observed in the purified wood. The results of X-ray analysis and tensile tests indicated that there were no significant differences among the sources in the cellulose crystallinity and Young’s modulus of the isolated microfibril aggregates in the dry state. These results suggest that the inherent characteristics of cellulose microfibril aggregates in the dry state are very similar regardless of plant sources and tissue functions.     

Atomic force microscopy and transmission electron microscopy of cellulose from Micrasterias denticulata; evidence for a chiral helical microfibril twist

Atomic force microscopy (AFM), tapping mode atomic force microscopy (TM-AFM) and transmission electron microscopy (TEM) have been used to image the cell wall, ultrathin sections of whole cells and cellulose microfibrils prepared from the green alga Micrasterias denticulata. Measurements of the microfibril dimensions are in agreement with earlier observations carried out by electron microscopy. Images at the molecular level of the surface of the microfibrils were obtained with AFM and show regular periodicities along the microfibril axis that correspond to the fibre and glucose repeat distances of cellulose. Twisted regions visible at intervals along the microfibrils dried down onto substrates were noted to be right-handed in over 100 observations by TEM, AFM and TM-AFM. This revised version was published online in August 2006 with corrections to the Cover Date.     

Cellulose microfibril orientation in onion (Allium cepa L.) epidermis studied by atomic force microscopy (AFM) and vibrational sum frequency generation (SFG) spectroscopy

Cellulose microfibril orientation in plant cell walls changes during cell expansion and development. The cellulose microfibril orientation in the abaxial epidermis of onion scales was studied by atomic force microscopy (AFM) and sum frequency generation (SFG) vibrational spectroscopy. Onion epidermal cells in all scales are elongated along the onion bulb axis. AFM images showed that cellulose microfibrils exposed at the innermost surface of the abaxial epidermis are oriented perpendicular to the bulb axis in the outer scales and more dispersed in the inner scales of onion bulb. SFG analyses can determine the orientation of cellulose microfibrils averaged over the entire thickness of the cell wall. We found that the average orientation of cellulose microfibrils inside onion abaxial epidermal cell walls as revealed by SFG is similar to the orientation observed at the innermost cell wall surface by AFM. The capability to determine the average orientation of cellulose microfibrils in intact cell walls will be useful to study how cellulose microfibril orientation is related to biomechanical properties and the growth mechanism of plant cell walls.     

Structural studies of bacterial cellulose through the solid-phase nitration and acetylation by CP/MAS <Superscript>13</Superscript>C NMR spectroscopy

The solid-phase nitration and acetylation processes of bacterial cellulose have been investigated mainly by CP/MAS ¹³C NMR spectroscopy to clarify the features of these reactions in relation to the characterization of the disordered component included in the microfibrils. CP/MAS ¹³C NMR spectra of bacterial and Valonia cellulose samples are markedly changed as the nitration progresses, in a similar way to the case of cotton linters previously reported; and the relative reactivity of the OH groups in the glucose residues is found to decrease in the order of O(6)H>O(2)H>O(3)H. Moreover, the nitration rate and mode greatly depend on the concentration of nitric acid in the reaction media. At dilute and medium concentrations, the O(6)H groups in the crystalline and disordered components are subjected to nitration at nearly the same rate, indicating that these two components are distributed almost at random in the entire region of each microfibril. The preferential penetration of nitric acid into each microfibril also occurs prior to nitration at the medium concentration, resulting in an increase in the mole fraction of the disordered component. In contrast, all OH groups undergo nitration very rapidly at the higher concentration, although nitration levels off to a certain extent for O(3)H groups. In solid-phase acetylation, no regio-selective reactivity is observed among the three kinds of OH groups, which may be due to the characteristic reaction that proceeds in a very thin layer between the acetylated and nonacetylated regions in each microfibril. The almost random distribution of the disordered component in the entire region of the microfibrils is also confirmed in this solid-phase acetylation. On the basis of these results, the mechanism of the solid-phase reactions and the microfibril structure are discussed.     

Vibrational sum-frequency-generation (SFG) spectroscopy study of the structural assembly of cellulose microfibrils in reaction woods

The cellulose microfibril assemblies in secondary cell walls of tension wood and compression wood were studied with vibrational sum frequency generation (SFG) spectroscopy. The tension wood contains the gelatinous layer with highly-crystalline and highly-aligned cellulose microfibrils. The SFG spectral features of tension wood changed depending on the azimuth angle between the polarization of the incident IR beam and the preferential alignment axis of the cellulose microfibrils. The SFG spectra of the compression wood did not show any dependence on the azimuth angle, implying that the overall orientation of cellulose microfibrils in compression wood is not highly aligned. Instead, the decrease of cellulose content in compression wood brought about larger separation between cellulose microfibrils, which was manifested as changes in CH₂/OH intensity ratio in SFG spectra. These results implied that SFG spectral features are sensitive to cellulose microfibril alignments and inter-fibrillar separations.     

Effect of alkaline treatment on cellulose supramolecular structure studied with combined confocal Raman spectroscopy and atomic force microscopy

The aim of this work was to investigate the morphological and structural changes associated with mercerization of cellulose fibres with combined confocal Raman and atomic force microscopy (AFM). During mercerization the alkali induces a change in polymorphic lattice from cellulose I to II. This was observed by confocal Raman spectroscopy from cellulose samples treated with 10, 15 and 25% aqueous sodium hydroxide solution. AFM images from the same samples illustrated that microfibrils were swollen and more granular in cellulose II than in cellulose I. Raman spectral images in plane and depth directions showed that the polymorphous cellulose structure was uniform throughout the cell wall, whereas the microfibril orientation varied between fibre cell wall layers. The changes in microfibril orientation on the sample surfaces were confirmed by AFM images measured from the same sample position.     

Influence of homogenization and drying on the thermal stability of microfibrillated cellulose

Homogenization has been used to release microfibrils from cellulose fibres to produce microfibrillated cellulose (MFC). Oven drying, atomization or freeze-drying were used to dry MFC. Morphological differences were observed linked to the compaction of the system and the formation of microfibril agglomerates. Thermal stability of the dried MFC, checked by TGA, decreased after homogenization and drying. Char level at the end of the pyrolysis was higher than for cellulose fibres. Derivative TGA (dTGA) showed a shoulder around 250 °C for the dried MFC. Volatile degradation product detection by FTIR spectroscopy (FTIR) coupled to TGA and DSC showed that the shoulder corresponds to expected dehydration reactions of the cellulose. Increasing the contacts between microfibril(s) (bundles) and agglomerates of the freeze-dried MFC by compression promoted dehydration reactions. Homogenization and drying modified the thermal properties of the MFC. No significant influence of freeze-drying kinetics on the thermal behaviour of the MFC was observed.     

Microfibril diameter in celery collenchyma cellulose: X-ray scattering and NMR evidence

Cellulose isolated from celery collenchyma is typical of the low-crystallinity celluloses that can be isolated from primary cell-walls of higher plants, except that it is oriented with high uniformity. The diameter of the microfibrils of celery collenchyma cellulose was estimated by three separate approaches: ¹³C NMR measurement of the ratio of surface to interior chains; estimation of the dimensions of the crystalline lattice from wide angle X-ray scattering (WAXS) measurements using the Scherrer equation; and the observation that microfibrils of this form of cellulose have the unusual property of packing into an irregular array from which small angle X-ray scattering (SAXS) shows features of both form and interference functions. The interference function contributing to the SAXS pattern implied a mean microfibril centre-to-centre distance of 3.6 nm, providing an upper limit for the diameter. However modelling of the scattering pattern from an irregular array of microfibrils showed that the observed scattering curve could be matched at a range of diameters down to 2.4 nm, with the intervening space more or less sparsely occupied by hemicellulose chains. The lateral extent of the crystalline lattice normal to the 200 plane was estimated as a minimum of 2.4 nm by WAXS through the Scherrer equation, and a diameter of 2.6 nm was implied by the surface: volume ratio determined by ¹³C NMR. The WAXS and NMR measurements both depended on the assumption that the surface chains were positioned within an extension of the crystalline lattice. The reliability of this assumption is uncertain. If the surface chains deviated from the lattice, both the WAXS and the NMR data would imply larger microfibril diameters within the range consistent with the SAXS pattern. The evidence presented is therefore all consistent with microfibril diameters from about 2.4 to 3.6 nm, larger than has previously been suggested for primary-wall cellulose. Some degree of aggregation may have occurred during the isolation of the cellulose, but the larger microfibril diameters within the range proposed are a consequence of the novel interpretation of the experimental data from WAXS and NMR and are consistent with previously published data if these are similarly interpreted.     

Parenchyma cell wall structure in twining stem of <Emphasis Type="Italic">Dioscorea balcanica</Emphasis>

Anatomical adaptation of liana plants includes structural changes in cell walls of different tissues: fibers, vessel elements and tracheids. However, the contribution of parenchyma cells to stem twining in liana plants is mostly unknown. The aim of this investigation is to determine changes in stem parenchyma cell walls that are correlated with the twinning process in liana plants. Parenchyma cell wall structure was studied on the stem cross sections of straight and twisted internodes of monocotyledonous liana Dioscorea balcanica, by different microscopy techniques: light microscopy, scanning electron microscopy, fluorescence detected linear dichroism microscopy and Fourier transform infrared microspectrometry. In addition, chemical analysis of the entire stem internodes was performed using photometric and chromatographic methods. Parenchyma cell walls of twisted D. balcanica internodes are characterized by: lower amounts of cellulose (obtained by FTIR microspectrometry) with different cellulose microfibril orientation (shown by Scanning electron microscopy), but no changes in “cellulose fibril order” (obtained by Differential polarization laser scanning microscopy); lower amounts of xyloglucan, higher amounts of xylan, higher amounts of lignin with modified organization—less condensed lignin (obtained by FTIR microspectrometry). At the same time, chemical analysis of the entire internodes did not show significant differences in lignin content and cell wall bound phenols related to stem twining, except for the presence of diferulate cross-links exclusively in twisted internodes. Our results indicate that adaptations to mechanical strain in D. balcanica stems involve modifications in parenchyma cell wall structure and chemistry, which provide decreased stiffness, higher strength and increased elasticity of twisted internodes.     

Mercerization and acid hydrolysis of bacterial cellulose

Structural changes in never- dried, disintegrated bacteria l cellulose by treatment with aqueous NaOH were examined by electron microscopy, X-ray diffractometry and acid hydrolysis behaviour and compared with those of cotton cellulose. The microfibril kept its fibrillar morphology after treatment with NaOH solutions of less than 9% (w/w), but changed into irregular aggregates when treated with NaOH above 12% (w/w), corresponding to the crystal conversion to cellulose II. The crystallinity of the resulting cellulose II was very low after a brief alkali treatment, but was increased significantly by elongated treatment (up to 10 days). In contrast, cotton cellulose was converted to cellulose II of fairly high crystallinity by alkali treatment of as little as 3 min duration, and the crystallinity did not change with longer treatments. The leveling-off degree of polymerization (LODP) of bacterial cellulose was decreased from 150 to 50 by 18% (w/w) NaOH treatment, while that of cotton linter decreased from 260 to 70. These characteristic differences between cotton linter cellulose and bacterial cellulose can be ascribed to a basic difference in microfibrillar organization in these materials: the microfibrils in cotton cellulose are in close contact with neighbouring microfibrils having opposite polarity, and in bacterial cellulose are isolated from each other and require chain folding to form the antiparallel cellulose II crystal     

Multiscale modeling of the elastic properties of natural fibers based on a generalized method of cells and laminate analogy approach

In this article, a new method based on a generalized method of cells and laminate analogy approach was used to predict the elastic properties of natural fibers. The elastic properties of cellulose crystals and amorphous cellulose were adopted to calculate the effective properties of microfibrils. A ten-layer antisymmetrical laminated structure was used to predict the effective properties of cell walls. The effects of the aspect ratio and volume fraction of cellulose crystal, the microfibril angle in the S2 layer and the lumen ratio of fiber on the axial Young’s moduli of natural fibers were analyzed in detail. The results show that the predicted properties of fibers are those of the cell fibers, and the final elastic properties of natural fibers can be obtained with the volume fractions of cell fibers as the corresponding conversion coefficients. The multiscale method is very effective in the predictions of the axial Young’s moduli of natural fibers.     

Accessibility and size of <Emphasis Type="Italic">Valonia</Emphasis> cellulose microfibril studied by combined deuteration/rehydrogenation and FTIR technique

We report an FTIR method to measure the accessibility and the size of cellulose microfibrils from the cell wall of Valonia ventricosa. This method is similar to the conventional deuteration technique for measuring the accessibility of cellulosic materials; however, the difference in our method is that the hydroxyl groups O2H, O3H, and O6H in the crystalline region were initially completely deuterated. The sample was then rehydrogenated by soaking in water at 25 °C, so that the OD groups on the surface were rehydrogenated. The ratio of OH to OD absorbance was used to calculate the number of surface vs. core cellulose chains in a microfibril. The obtained experimental ratio of 0.934 was consistent with the value calculated for a previously published 33 × 38 chain Valonia model (Sugiyama et al. 1984). The rehydrogenation process was further investigated by immersing the sample in water at elevated temperatures. At temperatures above 120 °C, rehydrogenation was more efficient, and the efficiency plots vs. rehydrogenation temperature showed two inflection. These points may correspond to the temperature where the cleavage of inter-chain hydrogen bonds and/or crystalline-phase transition would have been occurred.     

Microfibril orientation of the secondary cell wall in parenchyma cells of Phyllostachys edulis culms

The secondary cell wall of bamboo parenchyma cells is microfibril-based. However, understanding of the microfibril orientation of secondary cell walls in bamboo parenchyma cells is lacking. This study characterized the microfibril orientation of the sub-layers of the secondary cell wall in the parenchyma cells by field-emission environmental scanning electron microscopy and the microfibril angle of the ground parenchyma cell wall with X-ray diffraction. The microfibril orientation of tight-loose alternating layers of the secondary cell wall was in the opposite direction along the longitudinal axis. Near the parenchyma cells’ pit aperture, the microfibril orientation generally bypassed the pits and continued in a flow-like pattern. The mean microfibril angle of ground parenchyma cells was 63.3°. The average microfibril angle of adjacent sub-layers of the secondary cell wall was 60° and???65° in ground parenchyma cells, and 54° and???52° in vascular parenchyma cells. A structure model of microfibril orientation of parenchyma cell wall in moso bamboo was firstly constructed. The study provides insight into the anatomical structure of the parenchyma cell wall in the bamboo plant. Moreover, it provides a structural basis for further analysis of the mechanical properties of parenchyma cells.

     

Cellulose synthesizing terminal complexes in the ascidians

The structure of cellulose synthesizing terminal complexes (TCs) has been investigated in different species of ascidians. The linear-shaped TCs consisting of two types of membrane particles were found in four species belonging to two orders including previous data . A relationship between the length of TCs and the size of produced cellulose microfibrils was found within the same order. The density and localization patterns of TCs coincided with the arrangement of cellulose microfibrils in the innermost layer of the tunic, that is outer protective tissue and a major cellulosic component in the ascidians. This suggests that the shape and dimension of the microfibril bundle is influenced by an arrangement of TCs in the epidermal cell membrane.     

Deswelling of microfibril bundles in drying wood studied by small-angle neutron scattering and molecular dynamics

Cellulose - Structural changes of cellulose microfibrils and microfibril bundles in unmodified spruce cell wall due to drying in air were investigated using time-resolved small-angle neutron...     

The influence of thermo-hygro-mechanical treatment on the micro- and nanoscale architecture of wood cell walls using small- and wide-angle X-ray scattering

Tracking the changes of cellulose crystallites upon thermo-hygro-mechanical treatment is essential to understand the response of wood cell walls to steam and compression. In this paper the influence of Compression combined with Steam (CS) treatment on wood cellulose crystallites and pores structure of Chinese fir (Cunninghamia lanceolata) was studied under different steaming temperatures and compression ratios. Small-angle X-ray scattering and wide-angle X-ray scattering were used to investigate the changes of cellulose crystallites dimension, aspect ratio, fibril diameter distribution, non-crystalline fraction, the number of chains in each microfibril, as well as the fractal dimension and size of pores in response to CS treatment conditions. Results indicate that the crystallinity increased due to CS treatment, but did not show alteration with varying CS treatment conditions, i.e. seemed nearly unaffected by higher temperatures or compression ratio, both for earlywood and latewood. The cellulose crystallite diameter depended on processing parameters: it increased with increasing treatment temperature. No considerable differences were found for earlywood and latewood. We interpret our findings as a rearrangement of adjacent cellulose chains towards higher crystalline perfection attributing to the increase in crystallinity. The same effect allows a larger coherence length of crystalline order and therefore features an increasing cross-sectional dimension. In general we can state that the CS treatment leads to higher crystallinity and more perfectly arranged cellulose crystals, while it does not greatly affect the microfibril diameter but rather the amorphous regions of the microfibrils and the surrounding hemicellulose and lignin.     

Varietal difference in cellulose microfibril dimensions observed by infrared spectroscopy

We have previously reported a novel Fourier transform infrared (FTIR) method for evaluating both the accessibility and lateral dimensions of cellulose microfibrils. This method differs from conventional deuteration in that the OH groups in the crystalline region were initially completely deuterated. The samples were then rehydrogenated by immersing them in water at 25 °C, during which only the OD groups on the surface were rehydrogenated. The ratio of OD to OH groups measured for cellulose from various origins was used to estimate microfibril dimensions, which were compared with the data from X-ray diffractometry. The rehydrogenation process was further investigated by immersing the deuterated samples in water at elevated temperatures. The behavior of rehydrogenation under heat treatment was converted to observe the microfibril shape, which was in good agreement with the cross-sectional images obtained by diffraction contrast transmission electron microscopy techniques.     

Visualization of the nanoscale pattern of recently-deposited cellulose microfibrils and matrix materials in never-dried primary walls of the onion epidermis

For more than 10 years epidermal cell layers from onion scales have been used as a model system to study the relationship between cellulose orientation, cell growth and tissue mechanics. To bring such analyses to the nanoscale, we have developed a procedure for preparing epidermal peels of onion scales for atomic force microscopy to visualize the inner surface (closest to the plasma membrane) of the outer epidermal wall, with minimal disturbance and under conditions very close to the native state of the cell wall. The oriented, multilayer distribution of cellulose microfibrils, approximately ~3 nm wide, is readily observed over extended lengths, along with other features such as the distribution of matrix substances between and on top of microfibrils. The microfibril orientation and alignment appear more dispersed in younger scales compared with older scales, consistent with reported values for mechanical and growth anisotropy of whole epidermal sheets. These results open the door to future work to relate cell wall structure at the nm scale with larger-scale tissue properties such as growth and mechanical behaviors and the action of cell wall loosening agents to induce creep of primary cell walls.