Preparation and composition of mesophyll, epidermis and fibre cell walls from leaves of perennial ryegrass (lolium perenne) and italian ryegrass (lolium multiflorum)

Abstract

Cells of mesophyll, epidermis and residual fibrous material were obtained from leaves of Italian and perennial ryegrass harvested at different stages of maturity by mechanical disruption of leaf tissue. Mesophyll cells were selectively removed by filtration through 0.045 mm nylon mesh and remaining non-mesophyll cells centrifuged in metrizamide solutions (56-58% wt to vol.) of known density (1.308-1.329 gcm' at 5°C) to obtain a pure epidermis cell fraction and a residual fibre fraction. Whole mesophyll cells contributed 63-72%, epidermis 12-15% and the fibre fraction 15-24% to the total leaf dry matter. Fibre values were higher in late-cut samples. Cell walls were prepared from mesophyll and epidermis cells by disruption and washing to remove cell contents. Fibre cells were judged free of cell contents and received no further treatment. Examination of cell wall preparations by light and electron microscopy showed that both mesophyll and epidermis preparations were essentially free from contaminating material. Mesophyll cell walls were uniformly thin ~ (200 nm) while those of epidermis ranged from 200&3000nm at the outer face, thinning to 300nm or less at the inner surface. An electron-light layer (cuticle) of approximately 200 nm thickness was present covering the outer face of the epidermis. The fibre fraction largely consisted of sclerenchyma, but contained, in addition, other vascular cells, detached annular rings and heavily silicified leaf hairs. Analysis of cell walls accounted for 85-90% of dry matter. Cellulose was the major component of all cell walls examined (approximately 40% of dry matter) with xylose residues accounting for a further 11% of mesophyll, 13.5-17.5% of epidermis and 21-25% of fibre cell walls. Arabinose was low in fibre cells but was present in much higher proportions in mesophyll and epidermis walls. The ratio of arabinose to xylose was approximately 1:1.5 for mesophyll, 1:2.5 for epidermis and 1:7.0 for the fibre fraction. The molar ratio acetyl to xylose remained fairly constant at 1:4 regardless of the grass, cell type or maturity of the sample. The uronic acid content of epidermis was higher than that in other cell types and showed an increase with increasing maturity of the grass, reaching over 9 % in late-cut samples. Total phenolic material represented 2-3% of mesophyll and epidermis cell walls and 6 % of fibre walls. Ferulic acid alone was released from the primary cell walls by saponification and p-coumaric and ferulic acids from the secondary-thickened fibre walls. Crude protein values (Nx6.25) were high in mesophyll cell wall preparations and low in epidermis and fibre cell walls. Amino acid patterns were similar for both grasses and cell types but hydroxyproline was found in greater amounts in fibre cell walls than in either epidermis or mesophyll.

A 3-D Model of a Perennial Ryegrass Primary Cell Wall and Its Enzymatic Degradation

“…The 245 GAXs, 3 xyloglucans, 100 pectins and 35 (1,3;1,4)-β-glucans in the simulation contain respectively 9670, 4028, 2568 and 1028 monosaccharides, contributing 29.8%, 12.4%, 7.9% and 3.2% of the simulated wall. This is in good agreement with the analysis by Chesson and colleagues [20,[60][61][62] who found the composition of the perennial ryegrass MCW to be 46.6% cellulose, 29.8% GAX, 12.5% xyloglucans, 7.8% pectins and 3.2% (1,3;1,4)-β-glucans. Of the 1434 ferulates in the simulation, only 18 (1.2%) formed covalent bonds with another ferulate, and thus only nine diferulate cross-bridges were formed between the 245 GAXs.…”

“…Perma) [20,[60][61][62]. Our simulation was of a small volume of cell wall, involving segments of nine CMFs arranged in parallel (with the cellulose molecules 60 glucosyl residues long).…”

In‐vitro disappearance of carbohydrates, phenolic acids, and lignin from parenchyma and sclerenchyma cell walls isolated from cocksfoot

“…Parenchyma and sclerenchyma walls have very different chemical compositions (Gordon et al 1985;Grabber et a1 1991) and changes in the proportions of these cell types during digestion can lead to misleading results when heterogeneous preparations are used to study the relative IVD of cell-wall constituents. Analysis of isolated cell types may provide a more definitive means of studying the chemistry and digestion of forage fibre.…”

Characterisation of Lignin from Parenchyma and Sclerenchyma Cell Walls of the Maize Internode

“…If the nature of the lignin deposited in cell walls di †ers between cell wall layers, then this should be reÑected in a di †erence in composition of parenchyma and sclerenchyma at comparable stages of development. Sclerenchyma, parenchyma and various other cell types have been isolated from a variety of Gramineae by wholly mechanical means and cell wall preparations obtained (Gordon et al 1985 ;Grabber and Jung 1991 ;Lopez et al 1993). In this work advantage was taken of an earlier observation that cells at various stages of development could be conveniently isolated from a newly extended maize internode (Scobbie et al 1993).…”