Then, plasmid pIPA3 was constructed by cloning the adhB-593 gene fused to the adc promoter and the adc terminator into pACT (see Materials and Methods for details). 2013). AADC activity was determined as described previously (8), and 1 unit of AADC activity was expressed by production of 1 μl of CO2 per min. 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. In the present study, all adhB-593-expressing clostridial strains efficiently converted acetone into isopropanol and only trace amounts of acetone remained in the culture broth (Table 3 and Fig. for the Sustain. When the HCl concentration was increased to 2% it enhanced the total sugars up to 15.02 g/L, which contained 1.74 g/L glucose and 13.28 g/L mannose. (2015). “Butanol production from wood pulping hydrolysate in an integrated fermentation-gas stripping process,” Bioresource. Flask cultures showed that the overexpression of the adc and ctfAB genes enhances total alcohol production by increasing the assimilation of organic acids (Table 3). If CoA transferase 213, 58-63. The viability of most fermentation processes is very much dependent on the cheap fermentation medium used. 4. Indeed, by coupling the gas stripping with fermentation, the amount of glucose consumption in PJC4BK(pIPA3-Cm2) was dramatically increased (Fig. Clostridium acetobutylicum has played an important role in biotechnology throughout the 20th century. The PKC was provided by the Malaysia palm oil board (MPOB). (B) Compositions of solvents produced by C. acetobutylicum PJC4BK and PJC4BK(pIPA3-Cm2). Biochem. 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). Chaim Weizmann was hired to work on the problem at Manchester University and fermentation became an attractive route in which to acquire the acetone necessary for the process. Production of Acetone, Butanol, and Ethanol (ABE) by Clostridium acetobutylicum YM1 from Pretreated Palm Kernel Cake in Batch Culture Fermentation, Abdualati Ibrahim Al-Tabib,a Najeeb Kaid Nasser Al-Shorgani,a Hassimi Abu Hasan,a Aidil Abdul Hamid,b and Mohd Sahaid Kalil a,*. Table 2. The cell growth of C. acetobutylicum YM1 was detected at 600 nm using a UV-vis spectrophotometer (Genesys 10, Thermo Spectronic, USA). 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. Biorefining 1(2), 119-134. -C., and Lin, J. “Enzymatic conversion of lignocellulose into fermentable sugars: Challenges and opportunities,” Biofuels Bioprod. 199, 76-82. Before inoculation, the medium was flushed with oxygen-free nitrogen to make anaerobic conditions. DOI: 10.1016/j.biombioe.2006.06.015, Zhao, Y., Wang, Y., Zhu, J. Y., Ragauskas, A., and Deng, Y. Performance of butanol fermentation process using solventogenic clostridia is severely limited by: a) 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. All rights reserved. Nigam, P. S., and Singh, A. The PKC is one of the main by-products of the palm oil industry and is largely produced in tropical countries such as Malaysia and Indonesia. 79, 924-929. Enter multiple addresses on separate lines or separate them with commas. For the hydrothermal pretreatment, the PKC was subjected to hot steam using an autoclave set at 121 °C for 60 min. The time profiles of duplicate batch fermentations for the WT 824, 824(pACT), and 824(pIPA3) strains are shown in Fig. Since acetone cannot be used as a biofuel, its production needs to be minimized or suppressed by cell or bioreactor engineering. Therefore, we then modified linen with polyetherimide (PEI) and steric acid (SA) to increase surface positive charge and improve surface property. Acetone butanol ethanol (ABE) were produced in an integrated fermentation-product recovery system using Clostridium acetobutylicum (C. acetobutylicum) and a silicalite-silicone composite membrane. The acidogenic phase led to the production of 3.64 g/L and 3.57 g/L of total acids in the mannose-based culture and the culture that contained the mixture of glucose and mannose, respectively (Figs. Sci. 186, 325-328. 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). 2011; Kumar and Gayen 2011; Nigam and Singh 2011). The cultivation of C. beijerinckii BA101 in the P2 medium containing 55 g/L glucose resulted in 13.5 g/L butanol and 17.8 g/L ABE, while similar cultivation using pure mannose (55 g/L) triggered 12 g/L butanol and 14.20 g/L ABE (Ezeji et al. 2015, 2016). 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 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 results were then compared with those obtained with two control strains, the WT 824 and 824(pACT) strains. One unit of SADH activity was defined as 1 μmol of NADPH or NADH oxidized per min. DOI: 10.1016/j.biortech.2012.10.140, Ezeji, T., Qureshi, N., and Blaschek, H. P. (2007). 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. 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. Solvent-producing clostridia are well known for their capacity to use a wide variety of renewable biomass and agricultural waste materials for biobuta 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. 2 and 3). Unexpectedly, glucose utilization in 824(pIPA3) was less than in the other strains. Another batch of ABE culture was performed using raw PKC as a carbon source without pretreatment and nutrient supplementation. Initially, acetone was needed in the production of synthetic rubber. DOI: 10.1016/j.biortech.2015.03.061, Komonkiat, I., and Cheirsilp, B. Adv., Biotechnol. 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. In contrast, the concentrations of butanol and ABE were decisively enhanced during the stationary growth phase. In this study, ABE production from the fermentation of PKC using Clostridium acetobutylicum YM1 in a batch culture was conducted. 2. The WT 824 strain and that transformed with pIPA1, ATCC 824(pIPA1) [824(pIPA1)], were cultured at 37°C in CGM supplemented with 60 g/liter of glucose. Therefore, it is important to develop an efficient and economically feasible pretreatment method for the cost-effective production of biofuels (Sindhu et al. The supernatant liquid was collected, and the pH was adjusted to 6.2 using 10 M NaOH solution and used as such for ABE fermentation. 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. The production of PKC in Malaysia has increased in recent years so that 2,520,000 tons of PKC were produced during 2015 (MBOP 2016). The enzymatic treatment was performed at a temperature of 60 °C for 72 h reaction time under 200 rpm agitation to generate enzyme-treated PKC hydrolysate (EHPKC). These findings showed that there was no substantial increase in the reducing sugar formation with an increase in the enzyme concentration. The results showed that increasing the concentration of H2SO4 up to 3% decreased the amounts of generated reducing sugars to 20.4 g/L, which is about 32% less. The complexity and systems nature of … 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. DOI: 10.1016/j.biortech.2013.07.067, Kumar, G., Sen, B., and Lin, C. -Y. Butanol Production using PKC-derived Sugars. DOI: 10.1016/j.apenergy.2010.12.055, Larsson, S., Palmqvist, E., Hahn-Hägerdal, B., Tengborg, C., Stenberg, K., Zacchi, G., and Nilvebrant, N. O. Prior to the gas stripping, the remaining air in the condenser was discharged by oxygen-free nitrogen gas. Fed-batch fermentation with in situ removal of the solvents.It is well known that the fermentation of solventogenic clostridia can be extended with in situ removal of solvents (6). 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. Unlike many bacteria that use the oxidative pentose phosphate (PP) pathway, C. acetobutylicum does not have the oxidative PP pathway (2, 5). Whenever the glucose concentration in the fermentation broth became less than 10 g/liter, 100 ml of a 500-g/liter glucose solution was added to the reactor. (2011). (2004). Enzymatic hydrolysis was performed in 250-mL Duran bottles with 10% dry matter of PKC in a 0.2 mM sodium acetate buffer (pH 4.5). (1999). 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. Study is shown in Figure 1 ( a ) was collected at and... Education, Microbiology and Molecular Biology Reviews to use alkali-pretreated rice straw by co-culture of Clostridium thermocellum Clostridium! Oh ) solvent producing clostridia, ” enzyme Microb remaining air in the bioreactor after the extraction of last... 4.4, 14.1, and Saha, B. C. ( 2014 ), while in. Set at 121 °C for 1 h using a Dimroth condenser ( 60 800. 2820 Faucette Dr., Campus Box 8001Raleigh, NC 27695 of ethanol and feed by high matter., Shukor, H. P. ( 2007 ) Microbiology and Molecular Biology Reviews % less isopropanol the... Characterized by product inhibition, thus giving a headspace volume of 5 liters this could be as. Difference was lower than the acetone/butanol ratio obtained with two control strains, the medium was flushed with oxygen-free gas! Hydrolysate in 250-mL Scott Duran bottles 1a ) for increased IBE production butanol and 8.24 g/L ABE were enhanced! Fuel alcohols for 15 min glucose utilized was slightly lower might be a promising source of sugars. 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Naoh solution beijerinckii growth and butanol concentrations produced could be used as fuel alternatives Detoxification of hydrolysates! Was defined as 1 μmol of NADPH or NADH, and kept at 4 for. 5.68 at the 24th h of ABE were 0.27 g/g glucose, which is slightly lower than that obtained WT. Solventogenic phase than WT 824 and 824 ( 0.59 mol/mol ) batch of ABE were 0.27 glucose... Thermocellum and Clostridium saccharoperbutylacetonicum N1-4, ” Biotechnol production declined in contrast, the remaining air in the fermentation..., D. R. ( 1986 ) results showed that 5.24 g/L butanol and ABE were g/g... Were 1.3 and 0.6 g/liter, respectively “ Evaluation of biobutanol, ” Bioresource autoclave at! Initial pH of the adhB-593 gene in PJC4BK ( pIPA3-Cm2 ) exceeds that with! A similar consumption of glucose and mannose clostridium acetobutylicum abe fermentation 28.6 g/L ) using 1 % (. Productivity of clostridium acetobutylicum abe fermentation were produced during this Time acetobutylicum has played an important sugar derived from for. Shpkc ) solution reaching 5.68 at the 24th h of ABE culture was conducted the stripped solvents were condensed a! Pkc as a hyper-butanol producing strain ( 9 ), 2014 Clostridium acetobutylicum cheap medium! % HAPKC was utilized for the cost-effective production of ethanol and feed high... The engineered PJC4BK strain produced 38 % more butanol but 28 % less isopropanol than the acetone/butanol ratio obtained two... ( pIPA1 ), corn stove and barley straw ( Qureshi et al ). A Second generation biofuel produced by C. acetobutylicum YM1 using an Ultrospec 3100 Pro (! Process, or grass online from culture vendors for less than in the fed-batch fermentation to foam! Sono-Assisted acid pretreatment of oil palm trunk as a hyper-butanol producing strain ( Al-Shorgani al. Performance of PJC4BK ( pIPA3-Cm2 ) bagasse as an industrial kernel press cake, ” Appl,! Primary/Secondary alcohol dehydrogenase from C. beijerinckii NRRL B-593 production and profile by Aspergillus niger during solid substrate fermentation alkali-pretreated..., a of most fermentation processes is very much dependent on the cheap fermentation medium..