1, the acidogenic phase of the glucose-based culture was accompanied by a decrease in pH value (4.72) and an increase in the total acid concentration (3 g/L). This medium was used to prepare the inoculum of C. acetobutylicum YM1. For the ABE fermentation in continuous Clostridium acetobutylicum culture, this paper presents a kinetic model that includes the effects of key metabolic intermediates and enzymes as well as culture pH, product inhibition, and glucose inhibition. Technol. In the beginning of the ABE process, glucose, starch, whey permeate, and molasses were used as the traditional substrates, which are considered to be food-based substrates (García et al. The acetone–butanol–ethanol (ABE) fermentation was one of the largest biotechnological operations ever performed, being beaten in size only by the ethanolic fermentation. At 48 h after inoculation, WT 824 produced 3.8, 8.2, and 0.8 g/liter of acetone, butanol, and ethanol, respectively, from 40 g/liter of glucose (Table 3). Also, a higher yield (0.30 g/g glucose) and titer (20.4 g/liter) of the IBE mixture could be obtained by applying our strategy in the buk-deficient PJC4BK strain (Fig. In the past few years, many studies have focused on the use of lignocellulose materials, such as palm oil mill effluent (Kalil et al. ABE fermentation of hydrolysate from enzymatic saccharification of PKC. The hydrolysate obtained from SHPKC and HPPKC was used for ABE fermentation by C. acetobutylicum YM1 after adjustment of the pH to 6.2 with concentrated HCl. The PJC4BK(pIPA3-Cm2) strain was cultured at 37°C in a bioreactor containing 2 liters CGM supplemented with 80 g/liter glucose, and pH was maintained above 5.0. DOI: 10.1002/bit.21373, Fond, O., Matta-Ammouri, G., Petitdemange, H., and Engasser, J. M. (1985). DOI: 10.3923/pjbs.2003, Kiyoshi, K., Furukawa, M., Seyama, T., Kadokura, T., Nakazato, A., and Nakayama, S. (2015). Clostridium acetobutylicum were negatively charged surfaces during fermentation. The sulphuric acid-treated PKC (2% SAPKC) method produced the highest concentration of reducing sugars (30 g/L) compared with the other methods applied. Preliminary enzymatic hydrolysis of PKC was evaluated using mannanase enzyme with various enzyme loadings, namely 5% and 10% (w/wsubstrate), to determine the effect of enzyme loading on the amount of sugars released from pretreated-PKC. The pretreatment of lignocellulosic materials could constitute approximately 40% of the total biofuel production costs (Sindhu et al. DOI:10.1016/j.biotechadv.2009.06.002. One canonical example of such processes is acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum, during which cells convert carbon sources to organic acids that are later reassimilated to produce solvents as a strategy for cellular survival. “Pre-optimization of medium for biobutanol production by a new isolate of solvent-producing Clostridium,” BioResources 8(1), 1420-1430. In mixed sugar (glucose, xylose, galactose, and arabinose) fermentation, the culture preferred glucose and arabinose over galactose and xylose. One unit of SADH activity was defined as 1 μmol of NADPH or NADH oxidized per min. 24(3-4), 151-159 DOI: 10.1016/S0141-0229(98)00101-X, Long, S., Jones, D. T., and Woods, D. R. (1984). The second phase was the solvenogenic phase in which the acids were reassimilated, resulting in the production of solvents with an increase in pH value. This study showed that the pretreatment of PKC improved the content of fermentable sugars and subsequently enhanced the production of ABE by C. acetobutylicum YM1. When the concentration of H2SO4 was increased up to 3%, the amount of the total reducing sugar recovery decreased to 21.9 g/L, including 3.12 g/L glucose and 18.78 g/L mannose. (Chen et al. To date, however, this experiment has not been conducted using metabolically engineered clostridial strains. These findings showed that there was no substantial increase in the reducing sugar formation with an increase in the enzyme concentration. (2011) pretreated rice bran with 1% (v/v) H2SO4 and 1% (v/v) HCl for further utilization in the ABE process. Fig. 218, 257-264. -G. (2013). Energ. The butanol concentration obtained was 8.0 g/liter, which was comparable to that obtained with the wild type (8.2 g/liter) but less than that (8.9 g/liter) obtained with 824(pACT). Therefore, we then modified linen with polyetherimide (PEI) and steric acid (SA) to increase surface positive charge and improve surface property. Thus, this study aimed at developing an improved Clostridium acetobutylicum strain possessing enhanced fuel alcohol production capability. DOI: 10.1016/j.renene.2015.08.051, Cerveró, J. M., Skovgaard, P. A., Felby, C., Sørensen, H. R., and Jørgensen, H. (2010). Biochem. Microbial inoculum was prepared by transferring 1 mL of the spore suspension of C. acetobutylicum YM1 into 9 mL of the TYA medium, which was then heated for 1 min in boiling water, then cooled in iced water, and incubated for 1 to 2 days at 30 °C under anaerobic conditions. DOI: 10.1016/j.fuel.2015.05.073, Al-Shorgani, N. K. N., Isa, M. H. M., Yusoff, W. M. W., Kalil, M. S., and Hamid, A. The profile of ABE fermentations of glucose as the carbon source by C. acetobutylicum YM1: (a) solvents and OD; (b) sugar, pH, and acids concentration, Fig. Among different pretreatment methods, acid and alkali treatments are the most promising approaches that could enhance sugar recovery (Kumar et al. The concentrations of reducing sugars were measured by high-performance liquid chromatography (HPLC 12,000 Series; Agilent technologies, Palo Alto, CA, USA) using a SUPELCOGEL C-611 HPLC column (300 mm × 7.8 mm ID). The viability of most fermentation processes is very much dependent on the cheap fermentation medium used. 2011; Kumar and Gayen 2011; Nigam and Singh 2011). For comparison, the PKC was also pretreated with hydrochloric acid (HCl) by the addition of 100 g of PKC to 1 L of 1% and 2% HCl (v/v) solutions and was heated (121 °C and 45 min) to generate hydrochloric acid-pretreated PKC (HAPKC) sample. Briefly, the 7-liter bioreactor containing 1.8 liters CGM was used in the fed-batch fermentation to control foam without adding antifoam agent. After transformation, the recombinant PJC4BK strain harboring pIPA3-Cm2 was cultured at 37°C in a bioreactor containing 2 liter CGM supplemented with 80 g/liter glucose, and pH was controlled at above 5.0. Lignocellulosic biomass has been known to be a promising source of fermentable sugars. (1998). 15(2), 964-980. Journal of Microbiology & Biology Education, Microbiology and Molecular Biology Reviews. “Removal of fermentation inhibitors from alkaline peroxide pretreated and enzymatically hydrolyzed wheat straw: Production of butanol from hydrolysate using Clostridium beijerinckii in batch reactors,” Biomass Bioenerg. Table 3 shows the results of the acid pretreatment of PKC. This study approved that PKC is a suitable substrate for ABE fermentation. The pretreatment of de-oiled jatropha waste by acids also revealed that there was a positive relationship between the quantity of sugars released and the concentration of acids used for pretreatment (HCl (0.5% to 10%) and H2SO4 (0.5% to 5%)), so that the total sugar generated ranged from 1.4 g/L to 1.7 g/L and 1.4 g/L to 7.8 g/L using HCl and H2SO4, respectively (Kumar et al. 824(pIPA3) showed higher CoAT activities both in the acidogenic and in the solventogenic phase than WT 824. Solvent-producing clostridia are well known for their capacity to use a wide variety of renewable biomass and agricultural waste materials for biobuta Hydrolysate Obtained from the Acid Pretreated PKC and the Subsequent ABE Fermentation of the Hydrolysate by C. acetobutylicum YM1. Clostridium acetobutylicum YM1 was provided by the biotechnology lab, Department of Chemical and Process Engineering, Universiti Kebangsaan Malaysia (Bangi, Malaysia). Acetone was successfully converted to isopropanol by the expression of the adhB-593 gene in PJC4BK(pIPA3-Cm2) (Fig. 2010). The productivity and yield resulting from the fermentation of glucose were 0.02 g/L.h and 0.08 g/L, respectively. In the past decades, renewable lignocellulosic feedstocks have provided economic and sustainable options for cost-effective butanol production through microbial acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum [ 1, 2 ]. However, when the hydrolysate obtained from hydrochloric acid pretreatment was utilized, 4.93 g/L and 5.96 g/L of butanol and ABE were obtained, respectively. By combining in situ gas stripping with the fed-batch fermentation, very efficient IBE production was possible (35.4 g/liter within 45 h) (Fig. 6(8), 529-534. In this study, PKC was pretreated with a 1% NaOH (1% SHPKC) solution. The similar performance of EHPKC and 1% SAPKC to that of the control culture may have been due to the low concentrations of microbial inhibitors produced during the low severity pretreatment. of Human Soc. Lett. “Production of palm kernel cake for 2015,” Economic and Industry Development Division, (http://bepi.mpob.gov.my), (2016). We do not retain these email addresses. Hanai et al. The ABE production from glucose was similar to that produced from EHPKC and 1% SAPKC. Unexpectedly, glucose utilization in 824(pIPA3) was less than in the other strains. In the case of 824(pACT), acid reassimilation began earlier, and the pH began to rise when less than 25 g/liter of glucose was consumed. The maximum OD600 of 22.1 reached during gas-stripping fermentation was higher than that obtained from the batch fermentation (OD600, 17.1). Combust. ABE Fermentation of Glucose and Mannose by C. acetobutylicum YM1, Note: a The ratio of glucose and mannose is 1: 7. The ABE method devi… The results obtained from hydrochloric acid-pretreated PKC (HAPKC) showed that acid hydrolysis of PKC by 1% HCl recovered 5.69 g/L of total sugar (Table 3). The stripped solvents were condensed using a Dimroth condenser (60 by 800 mm) at −5°C. This culture was then subcultured in the TYA medium and incubated for 18 h to 20 h to be used as an inoculum source. Since 1-butanol is a promising biofuel, while acetone is not considered suitable as a fuel alternative, studies have been conducted to reduce acetone production. 4A). (2016a). The PKC was also pretreated with hydrogen peroxide as described by (Qureshi et al. Unlike many bacteria that use the oxidative pentose phosphate (PP) pathway, C. acetobutylicum does not have the oxidative PP pathway (2, 5). Jonsson, L. J., Palmqvist, E., Nilvebrant, N. O., and Hahn-Hagerdal, B. (A) Time profiles of the fed-batch fermentation of PJC4BK(pIPA3-Cm2). Similarly, the ABE fermentation was performed using 100 mL of PKC-derived hydrolysate in 250-mL Scott Duran bottles. The supplementation of SAPKC with the TYA (SAPKC as the only carbon source) medium enhanced the ABE production to 6.37 g/L (3.99 g/L butanol) compared with 5.72 g/L when SAPKC was used without the TYA supplement (Table 4). 135, 262-268. 2013). 4A) (132.9 g/liter). Subsequently, the mixture was centrifuged at 5000 rpm under room temperature for 15 min to remove the residual solids and the supernatant was kept at 4 °C for the reducing sugar analysis and ABE fermentation. (1999). The cell growth profile was monitored by measuring the optical density (OD 600) of clostridial cells. “Pretreatment and hydrolysis methods for recovery of fermentable sugars from de-oiled Jatropha waste,” Bioresour. Cell growth was monitored by measuring the OD600 using an Ultrospec 3100 Pro spectrophotometer (Amersham Biosciences, Uppsala, Sweden). Also, the complete consumption of glucose in our gas-stripping fermentation without strain degeneration suggests that the engineered strain developed here can be used for long-term fermentation. Since acetone cannot be used as a biofuel, its production needs to be minimized or suppressed by cell or bioreactor engineering. Bioeng. Copyright © 2020 American Society for Microbiology | Privacy Policy | Website feedback, Print ISSN: 0099-2240; Online ISSN: 1098-5336, Sign In to Email Alerts with your Email Address. 2. Biochem. DOI: 10.1016/j.biombioe.2008.04.009, Shukor, H., Abdeshahian, P., Al-Shorgani, N. K. N., Hamid, A. Zhen Chang†, Di Cai†, Chengyu Wang, Lun Li, Jiacheng Han, Peiyong Qin * and Zheng Wang * National Energy R&D Center for Biorefinery, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, PR China. Clostridium acetobutylicum. To study the growth of C. acetobutylicum YM1 in relation to the acidogenesis and solvenogenesis phases, three sets of ABE fermentations were performed in which C. acetobutylicum YM1 was grown in a TYA medium with glucose, mannose, and mixed sugars (glucose and mannose) at a ratio of 1:7. © 2020 NC State University. Thank you for sharing this Applied and Environmental Microbiology article. In this study, the PKC was pretreated with sulphuric acid at different concentrations ranging from 1% to 3% (v/v) for 45 min at 121 °C. The viability of most fermentation processes is very much dependent on the cheap fermentation medium used. Considering that the secondary alcohol dehydrogenase used in these studies is NADPH dependent (14), the insufficient conversion of acetone into isopropanol in engineered E. coli strains seems to be due to the cofactor imbalance. Jones, D. T., and Woods, D. R. (1986). (38), C. beijerinckii NRRL B-593 produced 2.2 and 3.7 g/liter of isopropanol and butanol, respectively, in the batch culture containing 60 g/liter of initial glucose. (2015). “Enzymatic conversion of lignocellulose into fermentable sugars: Challenges and opportunities,” Biofuels Bioprod. The low ABE and butanol concentrations produced could be attributed to the low sugar concentration in untreated PKC. It has been reported that CoAT might be a rate-limiting enzyme for acetone production (10, 41). Table 4. 2014), rice bran, and de-oiled rice bran (Al-Shorgani et al. The regular ratio of ABE solvents produced by C. acetobutylicum is 3:6:1 with 20 g/L being the maximum concentration achieved so far (Ranjan and Moholkar, 2009). This is due to the fact that the lower butanol concentration in the broth positively affected cell growth. The pretreatment results revealed that when H2SO4 and HCl were used, the concentrations of total sugar recovered from pretreated rice bran were 31.43 g/L and 9.7 g/L, respectively. The stripped solution was collected at 36 and 45 h, and its composition was analyzed. For the sulphuric acid pretreatment, 100 g of PKC was added to 1 L of 1%, 2%, and 3% (v/v) sulphuric acid solutions in 2-L Scott Duran bottles individually and was heated for approximately 45 min at 121 °C to generate sulphuric acid-pretreated PKC (SAPKC 1%, 2%, and 3%). Acetone butanol ethanol (ABE) were produced in an integrated fermentation-product recovery system using Clostridium acetobutylicum (C. acetobutylicum) and a silicalite-silicone composite membrane. It was then subjected to the different pretreatment methods, including acid pretreatment (sulphuric and hydrochloric acids), alkali pretreatment (sodium hydroxide and alkaline peroxide), enzymatic hydrolysis, and hydrothermal pretreatment to generate various samples labeled as SAPKC, HAPKC, SHPKC, HPPKC, EHPKC, and HPKC, respectively. The medium was sterilized at 121 °C for 15 min, and its initial pH was adjusted to 6.2. Biotech. To enhance the butanol production, the C. acetobutylicum PJC4BK strain, which is a buk-inactivated strain by Campbell-like integration and is known to be a better butanol producer than WT 824 (9, 12), was employed. Table 3 shows that the highest ABE yield obtained was from 2% and 3% SAPKC with the same value of 0.24 g/g. Results for the co-culture of the C. thermocellum and C. saccharoperbutylacetonicum strain N1-4 on alkali-pretreated rice straw hydrolysate showed that 5.5 g/L butanol was produced during 7 days of ABE fermentation (Kiyoshi et al. Cl (pH 7.5), 1 mM dithiothreitol, 0.2 mM NADPH or NADH, and 6.7 mM acetone (14). In another attempt, the PKC was pretreated using hot steam by autoclaving of PKC at 121 °C for 1 h. The pretreatment of PKC by hot steam showed that 1.10 g/L of reducing sugars were produced. 2015, 2016). The working volume of the reactor was maintained at 2 liters by adding sterilized distilled water to compensate for the loss of water due to the gas stripping. Contact information: a: Department of Chemical and Process Engineering, Faculty of Engineering and Built Environment, Universiti Kebangsaan Malaysia (UKM), 43600 UKM Bangi, Selangor, Malaysia; b: School of Biosciences and Biotechnology, Faculty of Sciences and Technology, Universiti Kebangsaan Malaysia (UKM), 43600 UKM Bangi, Selangor, Malaysia; * Corresponding author: sahaidster@gmail.com. Background: In acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum ATCC 824 using corn-based substrate, the solvents are generally produced at a ratio of 3:6:1 (A:B:E, w/w). For the sodium hydroxide pretreatment, PKC was added to a 1% NaOH solution to obtain a PCK loading of 10% (w/v). It was known as the ABE process, or acetone, butanol, ethanol process. Eng. There is a history regarding butanol production. Recently, the population dynamics of C. acetobutylicum was successfully investigated using flow cytometry (39), which might be useful for our future studies toward better understanding clostridial physiology and solventogenesis during gas-stripping-coupled fermentation. Among ABE, only butanol and ethanol can be used as fuel alternatives. Figures 1 to 3 show the growth profiles of C. acetobutylicum YM1 with variations in the solvents, organic acids, and culture pH measured using various sugars tested during 72 h of the batch ABE fermentations. The initial pH of the medium was adjusted to 6.2 by 10 M of NaOH solution. 37(1), 52-68. After enzymatic hydrolysis, the pH was adjusted to 6.2 and was used directly for ABE fermentation by C. acetobutylicum YM1. Before inoculation, the medium was flushed with oxygen-free nitrogen to make anaerobic conditions. Thus, enhanced flux from acetoacetyl-CoA to butyryl-CoA might have reduced the acetoacetyl-CoA pool and subsequently resulted in lower isopropanol production in PJC4BK(pIPA3-Cm2) than in 824(pIPA3). This performance of PJC4BK(pIPA3-Cm2) exceeds that of natural IBE producers. Photomicrographs of cytological changes in C. acetobutylicum during ABE fermentation. Until the end of the last century, the ABE fermentation from grain was operated in a few industrial scale plants. 2008). The alkali pretreatment of PKC was performed using sodium hydroxide 1% (w/v) and alkaline peroxide separately to generate sodium hydroxide-pretreated PKC (SHPKC 1%) and hydrogen peroxide-pretreated PKC (HPPKC) samples, respectively. To investigate the ability of C. acetobutylicum YM1 to ferment the mixture of glucose and mannose to butanol, an experiment was performed using the mixed sugars of glucose and mannose at a ratio of 1:7 (as in the H2SO4 pretreated hydrolysate) dissolved in the TYA medium. “Enhanced mannan-derived fermentable sugars of palm kernel cake by mannanase-catalyzed hydrolysis for production of biobutanol,” Bioresource Technol. , cost and efficiency are factors that limit the production of butanol and were! Potential of strain PJC4BK ( pIPA3-Cm2 ) strain was about 0.06 g/g solvents. Addition of ammonia solution carrier gas, and Pan, X, this study that! P262, ” Bioprocess Biosyst Developments in biobutanol production from the batch fermentation by ( Qureshi et.... Contrast, the ABE fermentation has a long history as an inoculum source ( see Discussion details. Authoritative coverage of both basic and clinical Microbiology decisively enhanced during the 72 h of ABE 11.4! 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