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Visualization of Miscanthus × giganteus cell wall deconstruction subjected to dilute acid pretreatment for enhanced enzymatic digestibility.

Ji Z, Zhang X, Ling Z, Zhou X, Ramaswamy S, Xu F - Biotechnol Biofuels (2015)

Bottom Line: DAP of M. × giganteus resulted in solubilization of arabinoxylan and cross-linking hydroxycinnamic acids in a temperature-dependent manner.The optimized pretreatment (1% H2SO4, 170°C for 30 min) resulted in significant enhancement in the saccharification efficiency (51.20%) of treated samples in 72 h, which amounted to 4.4-fold increase in sugar yield over untreated samples (11.80%).The consequently occurred changes in inner cell wall structure including damaging, increase of porosity and loss of mechanical resistance were also found to enhance enzyme access to cellulose and further sugar yield.

View Article: PubMed Central - PubMed

Affiliation: Beijing Key Laboratory of Lignocellulosic Chemistry, Beijing Forestry University, Beijing, 100083 China ; Ministry of Education Key Laboratory of Wooden Material Science and Application, Beijing Forestry University, Tsinghua East Road, Beijing, 100083 China.

ABSTRACT

Background: The natural recalcitrance of lignocellulosic plant cell walls resulting from complex arrangement and distribution of heterogeneous components impedes deconstruction of such cell walls. Dilute acid pretreatment (DAP) is an attractive method to overcome the recalcitrant barriers for rendering enzymatic conversion of polysaccharides. In this study, the internodes of Miscanthus × giganteus, a model bioenergy crop, were subjected to DAP to yield a range of samples with altered cell wall structure and chemistry. The consequent morphological and compositional changes and their possible impact on saccharification efficiency were comprehensively investigated. The use of a series of microscopic and microspectroscopic techniques including fluorescence microscopy (FM), transmission electron microscopy (TEM) and confocal Raman microscopy (CRM)) enabled correlative cell wall structural and chemical information to be obtained.

Results: DAP of M. × giganteus resulted in solubilization of arabinoxylan and cross-linking hydroxycinnamic acids in a temperature-dependent manner. The optimized pretreatment (1% H2SO4, 170°C for 30 min) resulted in significant enhancement in the saccharification efficiency (51.20%) of treated samples in 72 h, which amounted to 4.4-fold increase in sugar yield over untreated samples (11.80%). The remarkable improvement could be correlated to a sequence of changes occurring in plant cell walls due to their pretreatment-induced deconstruction, namely, loss in the matrix between neighboring cell walls, selective removal of hemicelluloses, redistribution of phenolic polymers and increased exposure of cellulose. The consequently occurred changes in inner cell wall structure including damaging, increase of porosity and loss of mechanical resistance were also found to enhance enzyme access to cellulose and further sugar yield.

Conclusions: DAP is a highly effective process for improving bioconversion of cellulose to glucose by breaking down the rigidity and resistance of cell walls. The combination of the most relevant microscopic and microanalytical techniques employed in this work provided information crucial for evaluating the influence of anatomical and compositional changes on enhanced enzymatic digestibility.

No MeSH data available.


Related in: MedlinePlus

Raman images of lignin redistribution within M. × giganteus cell walls upon dilute acid pretreatment. a Untreated; b pretreated M. × giganteus at 160°C, 0.5% H2SO4 for 15 min; c pretreated M. × giganteus at 170°C, 1% H2SO4 for 30 min. Sf sclerenchyma fibers, Par parenchyma, Pxv protoxylem vessel, Mxv metaxylem vessel, St sieve tube, Com companion cell.
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Fig7: Raman images of lignin redistribution within M. × giganteus cell walls upon dilute acid pretreatment. a Untreated; b pretreated M. × giganteus at 160°C, 0.5% H2SO4 for 15 min; c pretreated M. × giganteus at 170°C, 1% H2SO4 for 30 min. Sf sclerenchyma fibers, Par parenchyma, Pxv protoxylem vessel, Mxv metaxylem vessel, St sieve tube, Com companion cell.

Mentions: Raman images of lignin distribution were generated by integrating over the 1,575–1,620 cm−1 regions (Figure 7). The raw material had a heterogeneous distribution of lignin within various tissues, clearly displaying high intensity in Mxv, followed by Sf, and lowest in Pxv and Par (Figure 7a). Treatment of samples with aqueous acid caused redistribution of lignin, and the lignin content varied with pretreatment conditions. Based on the spectral analysis, the moderate pretreatment (160°C, 0.5% H2SO4 for 15 min) removed 5% lignin, while the harsher treatment (170°C, 1.0% H2SO4 for 30 min) resulted in greater lignin removal ca. 13%. Raman mapping technique helped visualization of lignin redistribution within specific tissues in selected portions of treated M. × giganteus. The lignin originally present in Pxv, Mxv and Sf walls largely resisted the moderate acid treatment, whereas the Par tissues were visibly delignified with evidence of a lower intensity at the bottom left of Raman images in Figure 7b. In comparison, severe pretreatment resulted in remarkable increase in the lignin signal intensity, especially in the Mxv (Figure 7c). One reason for this may be that with the removal of hemicelluloses during treatment, lignin was more exposed. Another possible explanation is that the dissolved lignin settled out from the bulk liquid and redeposited onto cell wall surfaces upon cooling after the pretreatment [38]. There are many factors that govern the extent of lignin relocalization following DAP, such as pretreatment severity, grass species and even the morphological regions tested. By scanning different areas, several vascular bundles were considerably conspicuous with an obvious reduction in lignin signal intensity at the same condition (see Additional file 1: Figure S4). Many investigations have suggested the effect of DAP on the fragmentation of lignin, usually leading to a slight delignification in biomass, the extent of which depends on the pretreatment severity [50–52]. Together, CRM imaging and TEM measurements provided more complete information on the removal, migration and relocalization of lignin resulting from DAP. Compared with the aforementioned compositional analysis, some differences in the residual lignin content may be attributed to the particle size of samples [53], from which thin sections were collected for Raman microspectroscopy. The intensity of lignin signal in Raman images was not directly related to the content of Klason lignin in samples. The primary focus of this section was on the dynamic relative distribution of lignin upon DAP, but not absolute lignin content. We suggest that the relocalization of lignin during DAP is as important as lignin removal in the context of glucan conversion, since both dramatically open up cell wall structures, leading to improved accessibility of cellulose to enzymes. Hydrothermal pretreatment has been reported to alter the role of lignin in biomass in terms of its association with pectins, arabinogalactans and xylans [34]. The work presented here supports that lignin content per se does not affect cellulose digestibility. Rather, the spatial distribution of lignin and its integration with other cell wall components appear to play a larger role.Figure 7


Visualization of Miscanthus × giganteus cell wall deconstruction subjected to dilute acid pretreatment for enhanced enzymatic digestibility.

Ji Z, Zhang X, Ling Z, Zhou X, Ramaswamy S, Xu F - Biotechnol Biofuels (2015)

Raman images of lignin redistribution within M. × giganteus cell walls upon dilute acid pretreatment. a Untreated; b pretreated M. × giganteus at 160°C, 0.5% H2SO4 for 15 min; c pretreated M. × giganteus at 170°C, 1% H2SO4 for 30 min. Sf sclerenchyma fibers, Par parenchyma, Pxv protoxylem vessel, Mxv metaxylem vessel, St sieve tube, Com companion cell.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC4513789&req=5

Fig7: Raman images of lignin redistribution within M. × giganteus cell walls upon dilute acid pretreatment. a Untreated; b pretreated M. × giganteus at 160°C, 0.5% H2SO4 for 15 min; c pretreated M. × giganteus at 170°C, 1% H2SO4 for 30 min. Sf sclerenchyma fibers, Par parenchyma, Pxv protoxylem vessel, Mxv metaxylem vessel, St sieve tube, Com companion cell.
Mentions: Raman images of lignin distribution were generated by integrating over the 1,575–1,620 cm−1 regions (Figure 7). The raw material had a heterogeneous distribution of lignin within various tissues, clearly displaying high intensity in Mxv, followed by Sf, and lowest in Pxv and Par (Figure 7a). Treatment of samples with aqueous acid caused redistribution of lignin, and the lignin content varied with pretreatment conditions. Based on the spectral analysis, the moderate pretreatment (160°C, 0.5% H2SO4 for 15 min) removed 5% lignin, while the harsher treatment (170°C, 1.0% H2SO4 for 30 min) resulted in greater lignin removal ca. 13%. Raman mapping technique helped visualization of lignin redistribution within specific tissues in selected portions of treated M. × giganteus. The lignin originally present in Pxv, Mxv and Sf walls largely resisted the moderate acid treatment, whereas the Par tissues were visibly delignified with evidence of a lower intensity at the bottom left of Raman images in Figure 7b. In comparison, severe pretreatment resulted in remarkable increase in the lignin signal intensity, especially in the Mxv (Figure 7c). One reason for this may be that with the removal of hemicelluloses during treatment, lignin was more exposed. Another possible explanation is that the dissolved lignin settled out from the bulk liquid and redeposited onto cell wall surfaces upon cooling after the pretreatment [38]. There are many factors that govern the extent of lignin relocalization following DAP, such as pretreatment severity, grass species and even the morphological regions tested. By scanning different areas, several vascular bundles were considerably conspicuous with an obvious reduction in lignin signal intensity at the same condition (see Additional file 1: Figure S4). Many investigations have suggested the effect of DAP on the fragmentation of lignin, usually leading to a slight delignification in biomass, the extent of which depends on the pretreatment severity [50–52]. Together, CRM imaging and TEM measurements provided more complete information on the removal, migration and relocalization of lignin resulting from DAP. Compared with the aforementioned compositional analysis, some differences in the residual lignin content may be attributed to the particle size of samples [53], from which thin sections were collected for Raman microspectroscopy. The intensity of lignin signal in Raman images was not directly related to the content of Klason lignin in samples. The primary focus of this section was on the dynamic relative distribution of lignin upon DAP, but not absolute lignin content. We suggest that the relocalization of lignin during DAP is as important as lignin removal in the context of glucan conversion, since both dramatically open up cell wall structures, leading to improved accessibility of cellulose to enzymes. Hydrothermal pretreatment has been reported to alter the role of lignin in biomass in terms of its association with pectins, arabinogalactans and xylans [34]. The work presented here supports that lignin content per se does not affect cellulose digestibility. Rather, the spatial distribution of lignin and its integration with other cell wall components appear to play a larger role.Figure 7

Bottom Line: DAP of M. × giganteus resulted in solubilization of arabinoxylan and cross-linking hydroxycinnamic acids in a temperature-dependent manner.The optimized pretreatment (1% H2SO4, 170°C for 30 min) resulted in significant enhancement in the saccharification efficiency (51.20%) of treated samples in 72 h, which amounted to 4.4-fold increase in sugar yield over untreated samples (11.80%).The consequently occurred changes in inner cell wall structure including damaging, increase of porosity and loss of mechanical resistance were also found to enhance enzyme access to cellulose and further sugar yield.

View Article: PubMed Central - PubMed

Affiliation: Beijing Key Laboratory of Lignocellulosic Chemistry, Beijing Forestry University, Beijing, 100083 China ; Ministry of Education Key Laboratory of Wooden Material Science and Application, Beijing Forestry University, Tsinghua East Road, Beijing, 100083 China.

ABSTRACT

Background: The natural recalcitrance of lignocellulosic plant cell walls resulting from complex arrangement and distribution of heterogeneous components impedes deconstruction of such cell walls. Dilute acid pretreatment (DAP) is an attractive method to overcome the recalcitrant barriers for rendering enzymatic conversion of polysaccharides. In this study, the internodes of Miscanthus × giganteus, a model bioenergy crop, were subjected to DAP to yield a range of samples with altered cell wall structure and chemistry. The consequent morphological and compositional changes and their possible impact on saccharification efficiency were comprehensively investigated. The use of a series of microscopic and microspectroscopic techniques including fluorescence microscopy (FM), transmission electron microscopy (TEM) and confocal Raman microscopy (CRM)) enabled correlative cell wall structural and chemical information to be obtained.

Results: DAP of M. × giganteus resulted in solubilization of arabinoxylan and cross-linking hydroxycinnamic acids in a temperature-dependent manner. The optimized pretreatment (1% H2SO4, 170°C for 30 min) resulted in significant enhancement in the saccharification efficiency (51.20%) of treated samples in 72 h, which amounted to 4.4-fold increase in sugar yield over untreated samples (11.80%). The remarkable improvement could be correlated to a sequence of changes occurring in plant cell walls due to their pretreatment-induced deconstruction, namely, loss in the matrix between neighboring cell walls, selective removal of hemicelluloses, redistribution of phenolic polymers and increased exposure of cellulose. The consequently occurred changes in inner cell wall structure including damaging, increase of porosity and loss of mechanical resistance were also found to enhance enzyme access to cellulose and further sugar yield.

Conclusions: DAP is a highly effective process for improving bioconversion of cellulose to glucose by breaking down the rigidity and resistance of cell walls. The combination of the most relevant microscopic and microanalytical techniques employed in this work provided information crucial for evaluating the influence of anatomical and compositional changes on enhanced enzymatic digestibility.

No MeSH data available.


Related in: MedlinePlus