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Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting.

Badhan A, Wang YX, Gruninger R, Patton D, Powlowski J, Tsang A, McAllister TA - Biomed Res Int (2015)

Bottom Line: Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design.Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes.The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

View Article: PubMed Central - PubMed

Affiliation: Agriculture and Agri-Food Canada, Lethbridge Research Centre, Lethbridge, AB, Canada T1H 4P4.

ABSTRACT
Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR) collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

No MeSH data available.


Related in: MedlinePlus

Glucose and xylose released as result of sequential ((a), (b)) and simultaneous ((c), (d)) hydrolysis of barley straw and alfalfa (biomass load: 2%, 30 mg of protein load/g of glucan in 5 mL reaction volume). Error bars (often invisible) represent SD of the mean (n = 8).
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fig4: Glucose and xylose released as result of sequential ((a), (b)) and simultaneous ((c), (d)) hydrolysis of barley straw and alfalfa (biomass load: 2%, 30 mg of protein load/g of glucan in 5 mL reaction volume). Error bars (often invisible) represent SD of the mean (n = 8).

Mentions: Predictions made by our micromodel were also validated in scale-up assays that used a solid load of 2% w/v of barley straw or alfalfa hay. The effect of enzyme prehydrolysis on the subsequent enhancement of cell wall hydrolysis was studied by sequential or simultaneous addition of recombinant enzymes to mixed rumen enzymes for barley straw and alfalfa hay. Results of hydrolysis of barley straw and alfalfa hay by mixed rumen enzymes after 48 h of prehydrolysis by endoglucanase EGL7A_THITE (50%) and ferulic acid esterase FAE 1a (50%) for barley straw and polygalacturonase PGA28A_ASPNG (50%) and arabinofuranosidase ABF54B_ASPNG (50%) for alfalfa hay confirmed that these mixtures increased the release of glucose and xylose (P < 0.05) as a result of prehydrolysis (Figures 4(a) and 4(b)). Supplementation of rumen mixed enzymes with endoglucanase EGL7A_THITE (50%) and ferulic acid esterase FAE 1a (50%) during the digestion of barley straw and polygalacturonase PGA28A_ASPNG (50%) and arabinofuranosidase ABF54B_ASPNG (50%) with alfalfa hay enhanced (P < 0.05) digestion as compared to mixed rumen enzymes alone (Figures 4(c) and 4(d)). A direct relationship was observed between xylan conversion (fraction of available xylan converted) and glucose conversion (fraction of available glucan conversion) during the hydrolysis of plant cell walls (Figure 5). However, a stronger correlation between xylan and glucan digestion with added auxiliary enzymes for optimized mixed rumen enzymes (Figure 5) suggested improved glucan conversion, perhaps due to better xylan saccharification.


Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting.

Badhan A, Wang YX, Gruninger R, Patton D, Powlowski J, Tsang A, McAllister TA - Biomed Res Int (2015)

Glucose and xylose released as result of sequential ((a), (b)) and simultaneous ((c), (d)) hydrolysis of barley straw and alfalfa (biomass load: 2%, 30 mg of protein load/g of glucan in 5 mL reaction volume). Error bars (often invisible) represent SD of the mean (n = 8).
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4477221&req=5

fig4: Glucose and xylose released as result of sequential ((a), (b)) and simultaneous ((c), (d)) hydrolysis of barley straw and alfalfa (biomass load: 2%, 30 mg of protein load/g of glucan in 5 mL reaction volume). Error bars (often invisible) represent SD of the mean (n = 8).
Mentions: Predictions made by our micromodel were also validated in scale-up assays that used a solid load of 2% w/v of barley straw or alfalfa hay. The effect of enzyme prehydrolysis on the subsequent enhancement of cell wall hydrolysis was studied by sequential or simultaneous addition of recombinant enzymes to mixed rumen enzymes for barley straw and alfalfa hay. Results of hydrolysis of barley straw and alfalfa hay by mixed rumen enzymes after 48 h of prehydrolysis by endoglucanase EGL7A_THITE (50%) and ferulic acid esterase FAE 1a (50%) for barley straw and polygalacturonase PGA28A_ASPNG (50%) and arabinofuranosidase ABF54B_ASPNG (50%) for alfalfa hay confirmed that these mixtures increased the release of glucose and xylose (P < 0.05) as a result of prehydrolysis (Figures 4(a) and 4(b)). Supplementation of rumen mixed enzymes with endoglucanase EGL7A_THITE (50%) and ferulic acid esterase FAE 1a (50%) during the digestion of barley straw and polygalacturonase PGA28A_ASPNG (50%) and arabinofuranosidase ABF54B_ASPNG (50%) with alfalfa hay enhanced (P < 0.05) digestion as compared to mixed rumen enzymes alone (Figures 4(c) and 4(d)). A direct relationship was observed between xylan conversion (fraction of available xylan converted) and glucose conversion (fraction of available glucan conversion) during the hydrolysis of plant cell walls (Figure 5). However, a stronger correlation between xylan and glucan digestion with added auxiliary enzymes for optimized mixed rumen enzymes (Figure 5) suggested improved glucan conversion, perhaps due to better xylan saccharification.

Bottom Line: Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design.Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes.The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

View Article: PubMed Central - PubMed

Affiliation: Agriculture and Agri-Food Canada, Lethbridge Research Centre, Lethbridge, AB, Canada T1H 4P4.

ABSTRACT
Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR) collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

No MeSH data available.


Related in: MedlinePlus