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

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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.


Butyric acid and total acid production by C. beijerinckii NCIMB 8052 in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde). CTL and SA represent the control and syringaldehyde-supplemented P2 medium. CTL_A (first arrow) and CTL_S (second arrow) represent acid production by C. beijerinckii grown in P2 medium at the peak of acidogenic and early solventogenic phase, respectively. SA_A (first arrow) and SA_S (second arrow) represent acid production by C. beijerinckii grown in syringaldehyde-supplemented P2 medium at the peak of acidogenic and early solventogenic phase, respectively. Second arrow shows marked decrease in acid concentration due to switch to solventogenic phase (control) and negligible decrease in acid concentration due to inability to switch to solventogenic phase (treatment). Note that gas chromatography recognizes and measures both acetic acid and acetate (component of P2 medium) as same compound. This fact explains why up to 4 g/L acid was measured at 0 h with an initial pH of 6.8
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Fig3: Butyric acid and total acid production by C. beijerinckii NCIMB 8052 in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde). CTL and SA represent the control and syringaldehyde-supplemented P2 medium. CTL_A (first arrow) and CTL_S (second arrow) represent acid production by C. beijerinckii grown in P2 medium at the peak of acidogenic and early solventogenic phase, respectively. SA_A (first arrow) and SA_S (second arrow) represent acid production by C. beijerinckii grown in syringaldehyde-supplemented P2 medium at the peak of acidogenic and early solventogenic phase, respectively. Second arrow shows marked decrease in acid concentration due to switch to solventogenic phase (control) and negligible decrease in acid concentration due to inability to switch to solventogenic phase (treatment). Note that gas chromatography recognizes and measures both acetic acid and acetate (component of P2 medium) as same compound. This fact explains why up to 4 g/L acid was measured at 0 h with an initial pH of 6.8

Mentions: Acetic and butyric acid production by C. beijerinckii during the acidogenic growth phase and their re-assimilation (solventogenic growth phase) are essential for butanol production and longevity of solventogenic Clostridium species during ABE fermentation (Ezeji et al. 2010). An initial drop in pH following an increase of acetic and butyric acid production during the acidogenic growth phase is evident at 12 h for both control and treatment fermentations (Fig. 2). While the pH profile of C. beijerinckii 8052 grown in P2 medium shows the typical biphasic fermentation (Ezeji et al. 2010), the pH profile of C. beijerinckii grown in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde) revealed a continuous decrease in pH falling to below pH 4.5 (Fig. 2). Acetic and butyric acid analysis showed that acid concentration in the fermentation medium reached 5.6 g/L (CTL_A) in 12 h prior to solventogenesis (CTL_S) during which a sharp decrease in acetic and butyric acid concentration occurred (Fig. 3; CTL_S) due to acid uptake and conversion to ABE. In contrast, C. beijerinckii grown in syringaldehyde-supplemented P2 medium accumulated up to 8.7 g/L acid (SA_A) in 24 h without marked decrease in acid concentration after 24 h (Fig. 3) due to an apparent failure of a “switch” from acidogenic to solventogenic stage culture, a phenomenon known as “acid crash”, which occasionally occurs in ABE fermentations. “Acid crash” occurs when the acid concentration in the fermentation broth exceeds the maximum tolerable limit or formic acid is produced, causing cessation of glucose uptake and rapid termination of solventogenesis (Maddox et al. 2000; Wang et al. 2011). It is worth mentioning that more than 3 g/L butyric acid, about seven times greater than that obtained from the control medium, accumulated in the fermentation medium during growth of C. beijerinckii in syringaldehyde-supplemented P2 medium (Fig. 3). Butyric acid inhibits microbial cells by disrupting cell membrane functions (Martin et al. 1983; Alsaker et al. 2009). Toxicity of butyric acid to solventogenic Clostridium species, in addition, increases as the pH decreases from 6.0 to 4.0 (Monot et al. 1984). Growth of C. beijerinckii in the syringaldehyde-supplemented P2 medium resulted in a pH below 4.5 (Fig. 2) and accumulation of toxic levels of butyric acid in the fermentation medium (Fig. 3). These data suggest that acid crash may be the real cause for the premature cessation of C. beijerinckii growth following 24-h fermentation and termination of the fermentation process.Fig. 2


Impact of syringaldehyde on the growth of Clostridium beijerinckii NCIMB 8052 and butanol production
Butyric acid and total acid production by C. beijerinckii NCIMB 8052 in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde). CTL and SA represent the control and syringaldehyde-supplemented P2 medium. CTL_A (first arrow) and CTL_S (second arrow) represent acid production by C. beijerinckii grown in P2 medium at the peak of acidogenic and early solventogenic phase, respectively. SA_A (first arrow) and SA_S (second arrow) represent acid production by C. beijerinckii grown in syringaldehyde-supplemented P2 medium at the peak of acidogenic and early solventogenic phase, respectively. Second arrow shows marked decrease in acid concentration due to switch to solventogenic phase (control) and negligible decrease in acid concentration due to inability to switch to solventogenic phase (treatment). Note that gas chromatography recognizes and measures both acetic acid and acetate (component of P2 medium) as same compound. This fact explains why up to 4 g/L acid was measured at 0 h with an initial pH of 6.8
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Related In: Results  -  Collection

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Fig3: Butyric acid and total acid production by C. beijerinckii NCIMB 8052 in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde). CTL and SA represent the control and syringaldehyde-supplemented P2 medium. CTL_A (first arrow) and CTL_S (second arrow) represent acid production by C. beijerinckii grown in P2 medium at the peak of acidogenic and early solventogenic phase, respectively. SA_A (first arrow) and SA_S (second arrow) represent acid production by C. beijerinckii grown in syringaldehyde-supplemented P2 medium at the peak of acidogenic and early solventogenic phase, respectively. Second arrow shows marked decrease in acid concentration due to switch to solventogenic phase (control) and negligible decrease in acid concentration due to inability to switch to solventogenic phase (treatment). Note that gas chromatography recognizes and measures both acetic acid and acetate (component of P2 medium) as same compound. This fact explains why up to 4 g/L acid was measured at 0 h with an initial pH of 6.8
Mentions: Acetic and butyric acid production by C. beijerinckii during the acidogenic growth phase and their re-assimilation (solventogenic growth phase) are essential for butanol production and longevity of solventogenic Clostridium species during ABE fermentation (Ezeji et al. 2010). An initial drop in pH following an increase of acetic and butyric acid production during the acidogenic growth phase is evident at 12 h for both control and treatment fermentations (Fig. 2). While the pH profile of C. beijerinckii 8052 grown in P2 medium shows the typical biphasic fermentation (Ezeji et al. 2010), the pH profile of C. beijerinckii grown in syringaldehyde-supplemented P2 medium (1.0 g/L syringaldehyde) revealed a continuous decrease in pH falling to below pH 4.5 (Fig. 2). Acetic and butyric acid analysis showed that acid concentration in the fermentation medium reached 5.6 g/L (CTL_A) in 12 h prior to solventogenesis (CTL_S) during which a sharp decrease in acetic and butyric acid concentration occurred (Fig. 3; CTL_S) due to acid uptake and conversion to ABE. In contrast, C. beijerinckii grown in syringaldehyde-supplemented P2 medium accumulated up to 8.7 g/L acid (SA_A) in 24 h without marked decrease in acid concentration after 24 h (Fig. 3) due to an apparent failure of a “switch” from acidogenic to solventogenic stage culture, a phenomenon known as “acid crash”, which occasionally occurs in ABE fermentations. “Acid crash” occurs when the acid concentration in the fermentation broth exceeds the maximum tolerable limit or formic acid is produced, causing cessation of glucose uptake and rapid termination of solventogenesis (Maddox et al. 2000; Wang et al. 2011). It is worth mentioning that more than 3 g/L butyric acid, about seven times greater than that obtained from the control medium, accumulated in the fermentation medium during growth of C. beijerinckii in syringaldehyde-supplemented P2 medium (Fig. 3). Butyric acid inhibits microbial cells by disrupting cell membrane functions (Martin et al. 1983; Alsaker et al. 2009). Toxicity of butyric acid to solventogenic Clostridium species, in addition, increases as the pH decreases from 6.0 to 4.0 (Monot et al. 1984). Growth of C. beijerinckii in the syringaldehyde-supplemented P2 medium resulted in a pH below 4.5 (Fig. 2) and accumulation of toxic levels of butyric acid in the fermentation medium (Fig. 3). These data suggest that acid crash may be the real cause for the premature cessation of C. beijerinckii growth following 24-h fermentation and termination of the fermentation process.Fig. 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.