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Impact of syringaldehyde on the growth of Clostridium beijerinckii NCIMB 8052 and butanol production

View Article: PubMed Central

ABSTRACT

While lignocellulosic biomass excels as a cheap, renewable resource for biofuel production, it does present some challenges such as generation of microbial inhibitory compounds. The mode of selective inhibition of acetone–butanol–ethanol (ABE) production (as opposed to cell growth) by syringaldehyde on Clostridium beijerinckii NCIMB 8052 was examined. C. beijerinckii 8052 grown in syringaldehyde-supplemented P2 medium had a comparable growth rate (μ = 0.34) at acidogenic growth phase to that of C. beijerinckii 8052 grown in control P2 medium (μ = 0.30). The addition of syringaldehyde into P2 medium inhibited solvent production by C. beijerinckii 8052 and increased butyric and acetic acid accumulation in the fermentation broth. Analysis of coenzyme A transferase (CoAT) using cell-free extracts of C. beijerinckii 8052 showed decreased expression and activity in the presence of syringaldehyde. These results indicate that C. beijerinckii 8052 CoAT is negatively affected by syringaldehyde and thus, hampers the ability of the microorganism to metabolize butyric and acetic acid for ABE production as evidenced by the accumulation of butyric and acetic acid in the fermentation broth.

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Representative 12% SDS-PAGE of C. beijerinckii 8052 protein extracts stained with Coomassie blue R-250. Lane 1: protein ladder; lanes 2 and 4: crude protein profile of C. beijerinckii during exponential growth phase (fermentation time, 12 h) in P2 medium (control); lanes 3 and 5: crude protein profile of C. beijerinckii during acidogenic growth phase (fermentation time, 12 h) in P2 medium containing syringaldehyde (1 g/L); lanes 6 and 7: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium; lanes 8 and 9: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium containing syringaldehyde (1 g/L). Test and control cultures were adjusted to an OD600 ≈1.0 at 12 and 24 h and cell pellets from 2 mL samples were lysed with tissue lyser (Qiagen, Hilden Germany) for protein extraction. Fewer proteins from solventogenic growing culture of C. beijerinckii were used for the SDS PAGE
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Fig5: Representative 12% SDS-PAGE of C. beijerinckii 8052 protein extracts stained with Coomassie blue R-250. Lane 1: protein ladder; lanes 2 and 4: crude protein profile of C. beijerinckii during exponential growth phase (fermentation time, 12 h) in P2 medium (control); lanes 3 and 5: crude protein profile of C. beijerinckii during acidogenic growth phase (fermentation time, 12 h) in P2 medium containing syringaldehyde (1 g/L); lanes 6 and 7: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium; lanes 8 and 9: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium containing syringaldehyde (1 g/L). Test and control cultures were adjusted to an OD600 ≈1.0 at 12 and 24 h and cell pellets from 2 mL samples were lysed with tissue lyser (Qiagen, Hilden Germany) for protein extraction. Fewer proteins from solventogenic growing culture of C. beijerinckii were used for the SDS PAGE

Mentions: Total protein from equal amounts (5.8 mg dry weight) of C. beijerinckii cells grown in P2 and syringaldehyde-supplemented P2 medium media were extracted during acidogenic and solventogenic growth phases as described above. In the presence of syringaldehyde, C. beijerinckii produced less protein, especially during stationary growth phase, when compared to growth in the absence of syringaldehyde as can be seen Table 2 and Fig. 5. Total protein levels of cell extracts of C. beijerinckii grown in control P2 and syringaldehyde-supplemented P2 media are the first evidence that syringaldehyde has a negative effect on protein concentration by C. beijerinckii strains. Following 24 h of fermentation, there is a 38% decrease in the amount of crude protein produced by C. beijerinckii grown in syringaldehyde-supplemented P2 medium compared to C. beijerinckii grown in control P2 medium (Table 2). At 36 h, there is a 52% decrease in total protein by C. beijerinckii grown in syringaldehyde-supplemented P2 medium compared to C. beijerinckii grown in control P2 medium. When total protein from equal amounts of C. beijerinckii cells grown in P2 and syringaldehyde-supplemented P2 medium media were extracted during acidogenic and solventogenic growth phases and applied onto SDS gel electrophoresis, band intensity of the crude proteins (Fig. 5) confirmed the results (decreased protein production by C. beijerinckii in the presence of syringaldehyde during solventogenic growth phase) obtained with Bradford protein assay method (Table 2). This indicates that syringaldehyde has a negative effect on protein concentration, including enzymes involved in ABE production pathway in C. beijerinckii strains. Interestingly, C. beijerinckii grown in syringaldehyde-supplemented P2 medium produced more protein than C. beijerinckii grown in control medium during initial stage (12 h) of fermentation (Table 2). The result may explain in part why C. beijerinckii had greater growth rate in the presence of syringaldehyde during the initial 24 h fermentation than the control fermentation.Table 2


Impact of syringaldehyde on the growth of Clostridium beijerinckii NCIMB 8052 and butanol production
Representative 12% SDS-PAGE of C. beijerinckii 8052 protein extracts stained with Coomassie blue R-250. Lane 1: protein ladder; lanes 2 and 4: crude protein profile of C. beijerinckii during exponential growth phase (fermentation time, 12 h) in P2 medium (control); lanes 3 and 5: crude protein profile of C. beijerinckii during acidogenic growth phase (fermentation time, 12 h) in P2 medium containing syringaldehyde (1 g/L); lanes 6 and 7: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium; lanes 8 and 9: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium containing syringaldehyde (1 g/L). Test and control cultures were adjusted to an OD600 ≈1.0 at 12 and 24 h and cell pellets from 2 mL samples were lysed with tissue lyser (Qiagen, Hilden Germany) for protein extraction. Fewer proteins from solventogenic growing culture of C. beijerinckii were used for the SDS PAGE
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Fig5: Representative 12% SDS-PAGE of C. beijerinckii 8052 protein extracts stained with Coomassie blue R-250. Lane 1: protein ladder; lanes 2 and 4: crude protein profile of C. beijerinckii during exponential growth phase (fermentation time, 12 h) in P2 medium (control); lanes 3 and 5: crude protein profile of C. beijerinckii during acidogenic growth phase (fermentation time, 12 h) in P2 medium containing syringaldehyde (1 g/L); lanes 6 and 7: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium; lanes 8 and 9: crude protein profile during solventogenic growth phase (fermentation time, 24 h) in P2 medium containing syringaldehyde (1 g/L). Test and control cultures were adjusted to an OD600 ≈1.0 at 12 and 24 h and cell pellets from 2 mL samples were lysed with tissue lyser (Qiagen, Hilden Germany) for protein extraction. Fewer proteins from solventogenic growing culture of C. beijerinckii were used for the SDS PAGE
Mentions: Total protein from equal amounts (5.8 mg dry weight) of C. beijerinckii cells grown in P2 and syringaldehyde-supplemented P2 medium media were extracted during acidogenic and solventogenic growth phases as described above. In the presence of syringaldehyde, C. beijerinckii produced less protein, especially during stationary growth phase, when compared to growth in the absence of syringaldehyde as can be seen Table 2 and Fig. 5. Total protein levels of cell extracts of C. beijerinckii grown in control P2 and syringaldehyde-supplemented P2 media are the first evidence that syringaldehyde has a negative effect on protein concentration by C. beijerinckii strains. Following 24 h of fermentation, there is a 38% decrease in the amount of crude protein produced by C. beijerinckii grown in syringaldehyde-supplemented P2 medium compared to C. beijerinckii grown in control P2 medium (Table 2). At 36 h, there is a 52% decrease in total protein by C. beijerinckii grown in syringaldehyde-supplemented P2 medium compared to C. beijerinckii grown in control P2 medium. When total protein from equal amounts of C. beijerinckii cells grown in P2 and syringaldehyde-supplemented P2 medium media were extracted during acidogenic and solventogenic growth phases and applied onto SDS gel electrophoresis, band intensity of the crude proteins (Fig. 5) confirmed the results (decreased protein production by C. beijerinckii in the presence of syringaldehyde during solventogenic growth phase) obtained with Bradford protein assay method (Table 2). This indicates that syringaldehyde has a negative effect on protein concentration, including enzymes involved in ABE production pathway in C. beijerinckii strains. Interestingly, C. beijerinckii grown in syringaldehyde-supplemented P2 medium produced more protein than C. beijerinckii grown in control medium during initial stage (12 h) of fermentation (Table 2). The result may explain in part why C. beijerinckii had greater growth rate in the presence of syringaldehyde during the initial 24 h fermentation than the control fermentation.Table 2

View Article: PubMed Central

ABSTRACT

While lignocellulosic biomass excels as a cheap, renewable resource for biofuel production, it does present some challenges such as generation of microbial inhibitory compounds. The mode of selective inhibition of acetone–butanol–ethanol (ABE) production (as opposed to cell growth) by syringaldehyde on Clostridium beijerinckii NCIMB 8052 was examined. C. beijerinckii 8052 grown in syringaldehyde-supplemented P2 medium had a comparable growth rate (μ = 0.34) at acidogenic growth phase to that of C. beijerinckii 8052 grown in control P2 medium (μ = 0.30). The addition of syringaldehyde into P2 medium inhibited solvent production by C. beijerinckii 8052 and increased butyric and acetic acid accumulation in the fermentation broth. Analysis of coenzyme A transferase (CoAT) using cell-free extracts of C. beijerinckii 8052 showed decreased expression and activity in the presence of syringaldehyde. These results indicate that C. beijerinckii 8052 CoAT is negatively affected by syringaldehyde and thus, hampers the ability of the microorganism to metabolize butyric and acetic acid for ABE production as evidenced by the accumulation of butyric and acetic acid in the fermentation broth.

No MeSH data available.