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In situ label-free imaging of hemicellulose in plant cell walls using stimulated Raman scattering microscopy

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ABSTRACT

Background: Plant hemicellulose (largely xylan) is an excellent feedstock for renewable energy production and second only to cellulose in abundance. Beyond a source of fermentable sugars, xylan constitutes a critical polymer in the plant cell wall, where its precise role in wall assembly, maturation, and deconstruction remains primarily hypothetical. Effective detection of xylan, particularly by in situ imaging of xylan in the presence of other biopolymers, would provide critical information for tackling the challenges of understanding the assembly and enhancing the liberation of xylan from plant materials.

Results: Raman-based imaging techniques, especially the highly sensitive stimulated Raman scattering (SRS) microscopy, have proven to be valuable tools for label-free imaging. However, due to the complex nature of plant materials, especially those same chemical groups shared between xylan and cellulose, the utility of specific Raman vibrational modes that are unique to xylan have been debated. Here, we report a novel approach based on combining spectroscopic analysis and chemical/enzymatic xylan removal from corn stover cell walls, to make progress in meeting this analytical challenge. We have identified several Raman peaks associated with xylan content in cell walls for label-free in situ imaging xylan in plant cell wall.

Conclusion: We demonstrated that xylan can be resolved from cellulose and lignin in situ using enzymatic digestion and label-free SRS microscopy in both 2D and 3D. We believe that this novel approach can be used to map xylan in plant cell walls and that this ability will enhance our understanding of the role played by xylan in cell wall biosynthesis and deconstruction.

Electronic supplementary material: The online version of this article (doi:10.1186/s13068-016-0669-9) contains supplementary material, which is available to authorized users.

No MeSH data available.


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Comparison of normalized Raman spectra. a Deacetylated corn stover (control). b Deacetylated corn stover with 55% cell wall xylan removed by xylanases. c Oat spelts xylan. d Birchwood xylan. e Xylo-oligomer (DP ranging from 2–7). fd-xylose
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Fig1: Comparison of normalized Raman spectra. a Deacetylated corn stover (control). b Deacetylated corn stover with 55% cell wall xylan removed by xylanases. c Oat spelts xylan. d Birchwood xylan. e Xylo-oligomer (DP ranging from 2–7). fd-xylose

Mentions: Figure 1 (top row) shows the Raman spectrum from the undigested deacetylated corn stover control containing 31% of xylan, as well as cellulose and lignin (Table 1). After xylanase digestion, 55% of the cell wall xylan was removed (the resulting Raman spectrum is also shown in Fig. 1). The Raman spectra from the xylan model compounds obtained from various sources, with more than 68% xylan content (Table 1), are also listed in Fig. 1 for comparison. Those xylan model compounds have a wide range of degrees of polymerization (DP) from d-xylose monomer to short oligomers with DP 2-7; as well as extracted xylan polymers with DP100–200.Fig. 1


In situ label-free imaging of hemicellulose in plant cell walls using stimulated Raman scattering microscopy
Comparison of normalized Raman spectra. a Deacetylated corn stover (control). b Deacetylated corn stover with 55% cell wall xylan removed by xylanases. c Oat spelts xylan. d Birchwood xylan. e Xylo-oligomer (DP ranging from 2–7). fd-xylose
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5120481&req=5

Fig1: Comparison of normalized Raman spectra. a Deacetylated corn stover (control). b Deacetylated corn stover with 55% cell wall xylan removed by xylanases. c Oat spelts xylan. d Birchwood xylan. e Xylo-oligomer (DP ranging from 2–7). fd-xylose
Mentions: Figure 1 (top row) shows the Raman spectrum from the undigested deacetylated corn stover control containing 31% of xylan, as well as cellulose and lignin (Table 1). After xylanase digestion, 55% of the cell wall xylan was removed (the resulting Raman spectrum is also shown in Fig. 1). The Raman spectra from the xylan model compounds obtained from various sources, with more than 68% xylan content (Table 1), are also listed in Fig. 1 for comparison. Those xylan model compounds have a wide range of degrees of polymerization (DP) from d-xylose monomer to short oligomers with DP 2-7; as well as extracted xylan polymers with DP100–200.Fig. 1

View Article: PubMed Central - PubMed

ABSTRACT

Background: Plant hemicellulose (largely xylan) is an excellent feedstock for renewable energy production and second only to cellulose in abundance. Beyond a source of fermentable sugars, xylan constitutes a critical polymer in the plant cell wall, where its precise role in wall assembly, maturation, and deconstruction remains primarily hypothetical. Effective detection of xylan, particularly by in situ imaging of xylan in the presence of other biopolymers, would provide critical information for tackling the challenges of understanding the assembly and enhancing the liberation of xylan from plant materials.

Results: Raman-based imaging techniques, especially the highly sensitive stimulated Raman scattering (SRS) microscopy, have proven to be valuable tools for label-free imaging. However, due to the complex nature of plant materials, especially those same chemical groups shared between xylan and cellulose, the utility of specific Raman vibrational modes that are unique to xylan have been debated. Here, we report a novel approach based on combining spectroscopic analysis and chemical/enzymatic xylan removal from corn stover cell walls, to make progress in meeting this analytical challenge. We have identified several Raman peaks associated with xylan content in cell walls for label-free in situ imaging xylan in plant cell wall.

Conclusion: We demonstrated that xylan can be resolved from cellulose and lignin in situ using enzymatic digestion and label-free SRS microscopy in both 2D and 3D. We believe that this novel approach can be used to map xylan in plant cell walls and that this ability will enhance our understanding of the role played by xylan in cell wall biosynthesis and deconstruction.

Electronic supplementary material: The online version of this article (doi:10.1186/s13068-016-0669-9) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus