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      Comprehensive Compositional Analysis of Plant Cell Walls (Lignocellulosic biomass) Part I: Lignin

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      Journal of Visualized Experiments : JoVE
      MyJove Corporation

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          Abstract

          The need for renewable, carbon neutral, and sustainable raw materials for industry and society has become one of the most pressing issues for the 21st century. This has rekindled interest in the use of plant products as industrial raw materials for the production of liquid fuels for transportation 1 and other products such as biocomposite materials 7. Plant biomass remains one of the greatest untapped reserves on the planet 4. It is mostly comprised of cell walls that are composed of energy rich polymers including cellulose, various hemicelluloses (matrix polysaccharides, and the polyphenol lignin 6 and thus sometimes termed lignocellulosics. However, plant cell walls have evolved to be recalcitrant to degradation as walls provide tensile strength to cells and the entire plants, ward off pathogens, and allow water to be transported throughout the plant; in the case of trees up to more the 100 m above ground level. Due to the various functions of walls, there is an immense structural diversity within the walls of different plant species and cell types within a single plant 4. Hence, depending of what crop species, crop variety, or plant tissue is used for a biorefinery, the processing steps for depolymerization by chemical/enzymatic processes and subsequent fermentation of the various sugars to liquid biofuels need to be adjusted and optimized. This fact underpins the need for a thorough characterization of plant biomass feedstocks. Here we describe a comprehensive analytical methodology that enables the determination of the composition of lignocellulosics and is amenable to a medium to high-throughput analysis. In this first part we focus on the analysis of the polyphenol lignin (Figure 1). The method starts of with preparing destarched cell wall material. The resulting lignocellulosics are then split up to determine its lignin content by acetylbromide solubilization 3, and its lignin composition in terms of its syringyl, guaiacyl- and p-hydroxyphenyl units 5. The protocol for analyzing the carbohydrates in lignocellulosic biomass including cellulose content and matrix polysaccharide composition is discussed in Part II 2.

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          Cell-wall carbohydrates and their modification as a resource for biofuels.

          Plant cell walls represent the most abundant renewable resource on this planet. Despite their great abundance, only 2% of this resource is currently used by humans. Hence, research into the feasibility of using plant cell walls in the production of cost-effective biofuels is desirable. The main bottleneck for using wall materials is the recalcitrance of walls to efficient degradation into fermentable sugars. Manipulation of the wall polysaccharide biosynthetic machinery or addition of wall structure-altering agents should make it possible to tailor wall composition and architecture to enhance sugar yields upon wall digestion for biofuel fermentation. Study of the biosynthetic machinery and its regulation is still in its infancy and represents a major scientific and technical research challenge. Of course, any change in wall structure to accommodate cost-efficient biofuel production may have detrimental effects on plant growth and development due to the diverse roles of walls in the life of a plant. However, the diversity and abundance of wall structures present in the plant kingdom gives hope that this challenge can be met.
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            Extraction and isolation of lignin for utilization as a standard to determine lignin concentration using the acetyl bromide spectrophotometric method.

            Lignin extracted with acidic dioxane was investigated as a possible standard for quantitatively determining lignin content in plant samples using the spectrophotometric method employing acetyl bromide. Acidic dioxane lignins were analyzed for carbohydrate, total protein, nitrobenzene oxidation products, and UV spectral characteristics. Total carbohydrate content of isolated lignins ranged from 2.21 to 5.70%, while protein ranged from 0.95 to 6.06% depending upon the plant source of the original cell wall sample. Nitrobenzene analysis indicated differences in the amount of guaiacyl and syringyl units making up the lignins, but this did not alter the UV spectrum of lignin solubilized in acetyl bromide. Regression equations developed for the acetyl bromide method using the isolated lignins for all the plant samples were similar to each other. Lignin values obtained by the acetyl bromide method were similar to the lignin values obtained as acid insoluble residues following a Klason lignin procedure.
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              Cell-wall carbohydrates and their modification as a resource for biofuels.

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                Author and article information

                Journal
                J Vis Exp
                JoVE
                Journal of Visualized Experiments : JoVE
                MyJove Corporation
                1940-087X
                2010
                11 March 2010
                11 March 2010
                : 37
                : 1745
                Affiliations
                Great Lakes Bioenergy Research Center, Michigan State University (MSU)
                Great Lakes Bioenergy Research Center and DOE-Plant Research Lab, Michigan State University (MSU)
                Author notes

                Correspondence to: Markus Pauly at paulymar@ 123456msu.edu

                Article
                1745
                10.3791/1745
                3144576
                20224547
                c6462c5f-5682-4f36-8bb8-101d7d805f57
                Copyright © 2010, Journal of Visualized Experiments

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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                Plant Biology

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